Synthetic studies and antibacterial activity of nucleobases and their N- and S-glucosides from 2-amino benzoic acid and its benzamido derivatives

Article abstract of DOI:10.13005/ojc/320528

A series of S-glucosides 11a-14a and their benzamido derivatives 11b-14b have been synthesized by reacting D-glucose with thiol groups of 5-(2′-aminophenylene)-1,3,4-oxadiazole-2-thioles 7(a,b), 5-(2′-aminophenylene)-1,3,4-thiadiazole-2-thiols 8(a,b), 5-(2′-aminophenylene)-1,2,4-triazole-3-thiols 9(a,b) and 5-(2′-aminophenylene)-4-N-amino-1,2,4-triazole-3-thiols 10(a,b). The thiols 7(a,b)-10(a,b) have been synthesized from hydrazides 3(a,b) which already been synthesized from 2-aminobenzoic acid and its benzamido derivative. All synthesized compounds were characterized by IR, UV,¹H- and¹³C- NMR. Nucleobases and a representative of S-glycoside were tested in vitro against the following microorganisms: two Gram-positive bacteria Staphylococcus aureus and Bacillus cereusand two Gram-negative bacteria Escherichia coli, Pseudomonas aeruginosa and they exhibited significant effects. Amykacine was used as positive standard.

Full text of DOI:10.13005/ojc/320528

ISSN: 0970-020 X
CODEN: OJCHEG
2016, Vol. 32, No. (5):
Pg. 2567-2576
ORIENTAL JOURNAL OF CHEMISTRY
An International Open Free Access, Peer Reviewed Research Journal
www.orientjchem.org
Synthetic Studies and Antibacterial Activity of Nucleobases
and their N- and S-Glucosides from 2-Amino Benzoic Acid and
its Benzamido Derivatives
SAMIA BENHAMMADI, SALIMA IRATEN and ADIL A. OTHMAN*
Laboratoire de Chimie Organique Bioactive, Département de Chimie Organique Industrielle, Faculte de
Chimie, Universite des Sciences et de la Technologie d’Oran- Mohamed Boudiaf- USTO-MB. B.P. 1505
ElM’naouer, Oran 31000, Algerie.
E.mail: adelaliothman@gmail.com
http://dx.doi.org/10.13005/ojc/320528
(Received: September 16, 2016; Accepted: October 16, 2016)
ABSTRACT
A series of S-glucosides 11a-14a and their benzamido derivatives 11b-14b have been
synthesized by reacting D-glucose with thiol groups of 5-(2’-aminophenylene)-1,3,4-oxadiazole-2-
thioles 7(a,b), 5-(2’-aminophenylene)-1,3,4-thiadiazole-2-thiols 8(a,b), 5-(2’-aminophenylene)-1,2,4-
triazole-3-thiols 9(a,b) and 5-(2’-aminophenylene)-4-N-amino-1,2,4-triazole-3-thiols 10(a,b).The thiols
7(a,b)-10(a,b) have been synthesized from hydrazides 3(a,b) which already been synthesized from
2-aminobenzoic acid and its benzamido derivative. All synthesized compounds were characterized
by IR, UV,1H- and13C- NMR. Nucleobases and a representative of S-glycoside were tested in vitro
against the following microorganisms: two Gram-positive bacteria Staphylococcus aureus and
Bacillus cereusand two Gram-negative bacteria Escherichia coli, Pseudomonas aeruginosa and
they exhibited significant effects. Amykacine was used as positive standard.
Keywords: Nucleobases, glycosides, benzamide derivatives, synthesis, antibacterial activity.
INTRODUCTION
or having different nucleobases like morpholine.2In
medicine several nucleosides analogues are used as
antiviral,3 anticancer,4 antibacterial5 and antifungal6
agents. Synthetic modified nucleosides designed
to pair in unusual ways with natural or unnatural
nucleobases have many potential applications in
biology; biotechnology and medicine have been
reported.7
Natural nucleosides and glycosides are
glycosylamines consisting of nucleobases (often
referred to as pyrimidine and purine derivatives).
They are bound to a ribose or deoxyribose sugar via a
beta-glycosidic linkage.1 In molecular biology, several
analogues of the sugar backbone exist e.g. locked
nucleic acid (LNA) and peptide nucleic acid (PNA)

2568
BENHAMMADI et al., Orient. J. Chem., Vol. 32(5), 2567-2576 (2016)
Since the discovery of nucleoside ribavirin8 of Oran-1, Algeria. The UV spectra were recorded
which made of triazole as its nucleobase, proved that on a ZUZI Split-Beam UV-Vis 4418PC (4418SPC)
it is not only possesses inhibitory activity against a spectrophotometer exhibited as (lmax in nm).The 1H
range of DNA and RNA viruses9 but also displays NMR and 13C NMR (250 MHz) spectra in DMSO-d6
antitumor activity in mice.10
exhibited as (d/ppm) were recorded at University of
Oran-1, Algeria.
The modern synthesis of nucleosides and
glycosides covered almost any kind of sugar-beta-
Microorganisms in this study supplied by
glycoside linkage and heterocyclic rings. Recently the university hospital of Oran and identified by
many C-, O-, N- and S-glycosides11-14 have received the laboratory of applied microbiology, University
considerable attention, because they are widely of Oran-1. The Mueller Hinton medium supplied by
employed as biological inhibitors, inducers, and (Difco).
ligands for affinity chromatography for carbohydrate
processing-enzymes and proteins15.
Synthesis
General procedures for esterification of 2-amino
Much attention has been focused on thiol benzoic acid.( 2-Amino-methyl and 2-amino-ethyl
derivatives of1,3,4- oxa, thia-diazoles, 1,2,4-triazole benzoates 2A and 2B)
and amino- 1,2,4-triazolefor their broad-spectrum Methyl esterification, catalysed by
activities, such as antitumor, anticonvulsant, Argil (Montmorillonite) to give 2-amino
antifungal16-19, herbicidal, and plant growth regulatory methylbenzoate2A
activities20.
An acid 1 ( 7g., 0.051 mole) dissolved in
methanol (200 ml) added to it dried argil – preheated
Till now, many of these diazole derivatives to 100°C for 1 hr (7.0g) and the mixture was refluxed
have been synthesized, and some of them have on water bath with an aid of magnetic stirring for
been patented for commercial uses 21.The synthesis 4 hr. After cooling, a saturated aqueous solution
and biological evaluation of N-glucosides16,22 and of NaOH (20ml) added and extracted twice with
C-glucosides containing 1,2,4-triazole have been dichloromethane (20 ml). The combined extracts
greatly emphasized, but only a few S-glucosides dried over anhydrous Na2SO4, filtered and evaporated
containing nucleobases of five membered to dryness to give 2-aminomethylbenzoate 2A as
heterocycles with three hetero atoms have been colourless syrup (5.1 g., 65%). Lit m.p. 24°C. IR,
reported.
n
max/ cm-1 3427 , 1729 (CO).
Recently, it has been reported that Ethyl esterification catalysed by H2SO4to give
4-benzamidobenzoic acid hydrazide derivatives 2-amino ethylbenzoate 2B
exhibited considerable in-vitro soluble epoxide
An acid 1 (5g. 0.02 mole) was dissolved
hydrolase inhibitory activity23,24. We also observed in ethanol (200 ml) added to it, drops of H2SO4 (5
that the benzamidobenzoic derivatives of all ml.). The mixture heated under reflux at 80°C for 8
synthesized intermediates had better solubility and hours. The reaction was monitored by TLC eluted
showed some different biological effects.
with chloroform/cyclohexane (6/4). The reaction
mixture was cooled down to room temperature and
neutralized by solid NaHCO3, filtered off and washed
with alcohol (2 ml.).The filtrate was dried over MgSO4
anhydrous, filtered and washed with alcohol (2ml),
EXPERIMENTAL
General
All reactions monitored by TLC, silica gel the combined filtrates were evaporated to dryness
F254, made by Merck, Germany.The melting points to give 2-amino ethylbenzoate 2B as colourless
measured with a BÜCHI 540 melting point apparatus syrup (1.8 g., 30%).Lit m.p. 13°-15°C. IR,nmax/ cm-1
and are uncorrected. The IR spectra exhibited as 3340 (NH2), 1670 (CO) , UV , lmax 192 nm. Loge
(wave number:v cm-1) were recorded using KBr discs 0.117; 194 nm. loge 0.119; 222nm. loge2.067; 248
in a JASCO V-530 spectrophotometer at University nm. loge0.763 ; 336 nm. loge0.542

2569
BENHAMMADI et al., Orient. J. Chem., Vol. 32(5), 2567-2576 (2016)
Benzoamidation of 2- amino benzoic acid 1H-NMR.dH 11.2 (s, 1H,NH),7.9-6.2 (m.4H,Har), 5.42
(2-Benzoamido benzoic acid (2C)
( s, H, NH), 4.1 ( d, 2H, NH2), 3.3 (SH).13C_NMR, 205
2-Amino benzoic acid 1(5.0 g, 0.033 mole), (C=S), 170.27 (N=C-SH), 169 ( N-C-O-), [132.76,
dissolved in chloroform (100 ml), benzoyl chloride 126.89, 117.30, 116.80, 77.44, 76.60 (Car)].
(50 ml) was added gradually, and the mixture was
cooled into an ice bath. Pyridine ( 5.0 ml) was added
2-Benzamido benzoic hydrazide (3b):White
dropwise with an aid of stirring.The reaction mixture crystalline (2.5 g., from 2D and 2.3 g., from 2E, 84%).
was refluxed at 110°C for 10 hr. Reaction was M.p. 195°C.IR,nmax/ cm-1 3342 (NH),2626 (SH),1630
1
monitored byTLC (CHCl3/cyclohexane, 6:4).Solvents (N-C=O).1610 (N-N-C=O), 1399 (C=S),UV, H and
were distilled under vacuum to give crystalline solid 13C-NMR are similar to above.
which on recrystallization from ethanol/hexane gave
a colourless crystalline 2-benzamido benzoic acid General procedure for preparation of 5-(2’-
(2C) (07.92 g, 90 %):M.p. 182-183°C. IR,nmax/ cm-1 benzamidophenylene)-1,3,4-oxadiazole-2-
3332 (NH), 1697( O-C=O), 1672 (N-C=O).
thiones 4(a,b) or thioles7(a,b)
The hydrazides 3a, (0.26 g.0.002 mole) or
Esterification of 2-benzoamido benzoic acid (2C), 3b,(1.0g., 0.004 mole) in an ethanolic solution of KOH
(2-Benzamidomethylbenzoate (2D)
85% (from KOH ,0.5 g. in ethanol 80 ml), added to
2-Benzoamido benzoic acid (2C) was them dropwise CS2 (2.5 ml for 3a and 10ml for 3b)
esterified with methanol and H2SO4–as above- to and the mixture was refluxed at 80°C for 7 to 22 hr.
give white crystalline 2- bezamidomethylbenzoate The bulk of the solvents removed under vacuum at
(2D), recrystallized from chloroform to give (4.5 g, 50°C, the remaining solid washed with iced diluted
85.6%). M.p.92-94°C. IR,nmax/ cm-1 3340 (NH), 1721 HCl in filter paper. The solid dissolved in a small
(O-C=O), 1672 (N-C=O).
amount of ethanol then left into the refrigerator to
give fine crystalline products:
2-Benzamidoethylbenzoate (2E)
2-Benzoamido benzoic acid (2C) was 5-(2’-aminophenylene)-1,3,4-oxadiazole-2-
esterified with ethanol and H2SO4 to give white thiones(4a or 7a)
crystalline 2- bezamidoethylbenzoate (2E) as white
White crystalline ( 0.233 g, 70%), m.p.
crystalline, recrystallized from chloroform to give 210°C .IR,nmax/ cm-1 3410.2 strong and broad
(3.7 g.,66 %): M.p.94-98°C. IR,nmax/ cm-1 3340 (NH), (free and bonded NH), 2626 (SH), 1632 (C=N),
1720 (O-C=O), 1672 (N-C=O).
1403 (C=S).UV lmax 240, 350 nm. loge 6.0, 6.0
respectively.NMR.dH 9.7 (s, 1H, NH oxadiazole ring),7.1-6.65
General procedure for preparation of hydrazides (m, 4H, Har) ,6.2 (s,2H,NH2), 3.4 (s, 1H,SH).13C-NMR
3a and 3b
The esters 2(A-D) (3 g.) dissolved in ethanol 132.16, 129.8,126.5, 116.94, (Car).
(80 ml), added to it dropwise a hydrazine hydrate
(d/ppm):205.54(C=S ), 162.11(C=N), 164.56, 155.4,
64% (5ml.) and the mixtures were heated under 5-(2’-benzoamidophenylene)-1,3,4-oxadiazole-2-
reflux on a water bath 90°C (20 hr. for 2(A-C) and thiones(4b or 7b)
12 hr for 2D. The reaction monitored by TLC. The
White crystalline (0.90 g, 78%), m.p.278°C.
solvents were evaporated under reduced pressure IR,nmax/ cm-1,3333 strong and broad (free and bonded
to give white solids which were washed with a small NH), 2626 (SH), 1502 (C=O), 1382 (C=S).UV lmax
1
quantity of ether ( few mls) to give corresponding 242, 348 nm. loge 6.0, 6.0 respectively. H-NMR
crystalline products
(d/ppm), 10.2 (s, 1H, NH oxadiazole ring), 7.8-6.30 (m, 9H,
Har) , 6.2 (s,H,NH), 3.4 (s, 1H,SH).13C-NMR (d/ppm):
2-aminobenzoichydrazide (3a): White 198.56(C=S ), 164.34(C=N), 166.36-,115.23(Car)
crystalline (2.4 g. from 2A and 2.7 g. from 2B, 90%):
M.p.119-122°C; IR,lmax/ cm-1 3333 (NH), 2626 (SH), General procedure for preparation of
1631 (N-CO), 1610 (N-N-C=O), 1382 (C=S). UV 2-aminophenylenethiosemicarbazides 5a and
,lmax 192, 195, 208, 214, 216, 248, 326, loge 0.539, 5b
0.617, 2.814, 3.077, 3.039, 1.451, 0.68 respectively.
The hydrazides 3a (2.0 g., 0.013 mole) and

BENHAMMADI et al., Orient. J. Chem., Vol. 32(5), 2567-2576 (2016)
2570
3b (1.0 g., 0.003 mole) dissolved in ethanol (200 ml
for 3a and 80 ml for 3b) added to it NH4SCN (1.7 g.,
for 3a and 0.85 g., for 3b). HCl (15 ml for 3a and 10
ml for 3b were added dropwise, refluxed at 80°C for
14 hr. Bulk of ethanol was removed by vacuum, the
thiosemicarbazides 5a and 5b were precipitated,
filtered off and recrystallized from methanol to
give:
g.,78%),m.p. 267°C. IR,nmax/ cm-1 3290 (NH),
1642(N-C=O), 1402 (C=S). NMR.dH 10.2 (s, 1H, NH
oxadiazole ring), 7.3-6.85 (m, 4H, Har) , 6.5 (s,2H,NH2), 3.2
(s, 1H,SH).13C-NMR (d/ppm): 198.88(C=S ), 165.88
(C=O), 162.45 (C=N),[160.65- 110.24, (Car)].
General procedure for preparation of 1,2,4-
triazoles 9a and 9b
2-Aminophenylenethiosemicarbazide(5a). 2.4 g.,
86%. M.p. 180°C.IR,nmax/ cm-1 3290 (NH), 1631
(C=O), 1404 (C=S).
2-Benzamidophenylenethiosemicarbazide(5b).0.98
g., 80%. M.p. 222°C.IR,nmax/ cm-1 3233 (NH), 1620
(C=O), 1404 (C=S).
The thiosemicarbazides 6a (1.0g., 0.005
mole) dissolved in water (100 ml). An aqueous
solution of NaOH (0.42 g.NaOH, in 30 ml of H2O)
was added and the mixture wax refluxed for 10 hr.
Water was evaporated under vacuum, solid formed,
recrystallized from methanol to give brownish
crystalline:
General procedure for preparation of thiadiazoles
8a and 8b
5-(2’-Aminophenylene)-1H-1,2,4-triazole-
3-thiol(9a): (0.86 g., 89%). M.p. 179°C. IR,nmax/ cm-1
, 3426 (NH, free), 3223 (NH, bonded), 2600 (SH),
1631 (C=N), 1403(C=S). 1H-NMR.dH 9.5 (s, 1H, NH
triazole), 7.9-7.3 (m, 4H, Har) , 3.4 (s, 1H,SH).13C-NMR
(d/ppm): 204.60(C=S ), 147.9(C=Ntriazole),[130.56-
116.94, (Car)].
The thiosemicarbazides 5a and 5b( 1.0g.)
were dissolved in absolute ethanol (100 ml), KOH
(0.5 g) was added with an aid of strring, followed by
addition of CS2 ( 1.0ml).The mixture was refluxed at
a temperature 75-80°C for 5 hr., cooled and acidified
with HCl to pH=1to give :
The thiosemicarbazides 6b (1.0g., 0.003
mole) dissolved in water (100 ml). An aqueous
solution of KOH (0.53g.KOH, in 20 ml of H2O)
was added and the mixture wax refluxed for 8
hr. Water was evaporated under vacuum, solid
formed, recrystallized from ethanol to give brownish
crystalline:
5-(2’-Aminophenylene)-1,3,4-thiadiazole-2-
thiols(8a):white crystalline, (0.8 g., 80%),m.p.167°C.
IR,nmax/ cm-1 3453 (NH), 1632 (C=N), 1403 (C=S).
NMR.dH 9.7 (s, 1H, NH oxadiazole ring), 7.1-6.65 (m, 4H,
Hphenyl) , 6.2 (s,2H,NH2), 3.4 (s, 1H,SH).13C-NMR
(d/ppm): 205.54 (C=S ), 162.11 (C=N), 164.56,
155.4, 132.16, 129.8,126.5, 116.94, (Car).
5-(2’-Benzamidophenylene)-1H-1,2,4-
triazole-3-thiol(9b): (0.86 g., 91%). M.p. 252°C.
IR, nmax/ cm-1 . 3350 (NH, free and bonded), 2604
(SH), 1664 (N-C=O),1631 (C=N),1402 (C=S).
5-(2’-Benzamidophenylene)-1,3,4-
thiadiazole-2-thiols(8b): white crystalline,(O.78
R
H
H
R
NH2
O
O
O
N
N
i--iv
v
OH
O-R'
NH-NH2
1
3a,R=H
3b,R=COph
2A,R=H;R'=CH3
2B,R=H;R'=C2H5
2C,R=COph;R'=H
2D,R=COph;R'=CH3
2E,R=COph;R'=C2H5
i-CH3OH,H2SO 4 ; ii-C2H5OH,H2SO 4;iii-C2H5OH;Argile; iv-,PhCOCl;iv-NH2NH 2.H 2O
Scheme1: Synthesis of hydrazides3a and 3b

2571
BENHAMMADI et al., Orient. J. Chem., Vol. 32(5), 2567-2576 (2016)
NMR.dH 10.2 (s, 1H, NH triazole), 7.6-6.68 (m, 9H, Har), (C=N).NMR.dH 8.9 (s, 1H, NHtriazole), 7.6-6.8 (m, 4H,
3.5 (s, 1H,SH).13C-NMR (d/ppm): 199.60(C=S ), Har) , 6.52 (s,2H,NH2), 3.4 (s, 1H,SH).13C-NMR (d/
165( C=O),136.8(C=Ntriazole), [132.44-112.94, (Car)].
ppm): 147.9(C=Ntriazole),[130.56-116.94, (Car)].
General procedure for preparation of 4-amino-1,
2, 4-triazoles 10a and 10b
5-(2’-Benzoamidophenylene)-4-N-amino-
1,2,4-triazole-3-thiol(10b) (1.06 g., 80%), m.p.
The hydrazides 3a and 3b (1.0 g., 0.007 225°C.IR,nmax/ cm-1 3290 (NH), 2630 (SH), 1643
mole and 0.003 mole respectively) dissolved in (N-C=O), 1624( C=N), 1398 (C=S).1H-NMR.dH
methanol (50 ml), KOH (1.86 g) was added followed 9.6 (s, 1H, NH triazole), 7.8-6.35 (m, 9H, Har), 3.2
by dropwise addition of CS2 (5 ml).The mixture was (s, 1H,SH).13C-NMR (d/ppm): 202.40 (C=S ), 163
stirred at room temperature for 15 hr. Diethyl ether ( C=O),135.0(C=Ntriazole),[133.44-110.5, (Car)].
(30 ml) added and stirring continued for further
1.0 hr to give a white ppt of salt 6a and 6b. Salts General procedure for preparation of
6(a,b)–individually-, without further identification, S-nucleosides 11(a,b)- 14(a,b)
were added to it hydrazine hydrate 64% (2 ml) and
Thiols 7(a,b)-10(a,b) ( 0.01 mole) and
the reaction mixture was refluxed at 80°C for 8 hr. D-glucose ( 0.01 mole) were dissolved in ethanol
Cooled to 5°C and acidified with HCl to pH=1 to give (100 ml) added to it HCl ( 8 ml) and the mixture was
colourless crystalline, recrystallized from ethanol :
refluxed at 80°C for 5-8 hr.Proceeding of the reaction
was monitored by tlc. Volatile solvent removed by
5-(2’-Aminophenylene)-4-N-amino-1,2,4- vacuum, the syrup or the solids remain were washed
triazole-3-thiol(10a)(1.27g., 93%),m.p.160°C;IR, with ether to give:
n
max/ cm-1 3423 (NH), 3063 (CHar), 2473 (SH), 1622
3(a,b)
i
iii
Route A
Route C
ii
Route B
O
O
R
O
NH-NH-CS-O-,K +
S
R
NH-NH-CS-NH2
R
NH
N
6(a,b)
5(a,b)
4(a,b)
v
iv
vi
N
N
N
N
N
O
N
N
N
R
SH
SH
N
SH
R
R
R
N
H
SH
S
H2N
10(a,b)
9(a,b)
7(a,b)
8(a,b)
H
H
COPh
H
N
Ra =
Rb =
N
;
i: CS2, KOH, Reflux; ii: NH4SCN; iii: KOH,CS2,Room temp; iv: KOH, CS2, EtOH, HCl; : NaOH, H2O; vi:
NH2NH2, KOH
Scheme2: Synthesis of nucleobases 7(a, b)-10(a, b) from hydrazides 3(a,b)

BENHAMMADI et al., Orient. J. Chem., Vol. 32(5), 2567-2576 (2016)
2572
5-(2’-Aminophenylene)-1,3,4-oxadiazole-2-
S-glucoside (11a), yellow syrup. IR, nmax/ cm-1 3423
(OH), 3290 (NH), 3067 (CH), 2998 (CHar),1630
(C=N).RMN, 8.3 (s,1H,NH), 8-7.5 (m, 4H, Har),4.5(m,
4H,O-H); 3.7(m,8H, C-H),3.2(d,2H, H2-C-O).13C-
NMR (d/ppm):167( C=N), 140-120(Car), 78-72
5-(2’-Aminophenylene)-1,3,4-triazole-2-S-
glucoside (13a) : Recrystallized from ethanol: M.p.
77°C.IR, nmax/ cm-3423 (OH), 3290 (NH), 3067 (CH),
2998 (CHar), 1630 (C=N).1H-NMR, 8.4 (s,1H,NH),
7.5-7(m, 4H, Har),4.5(m, 4H,O-H);3.6(m,8H,H-C-O),
3.3(d,2H, H2C-O).13C-NMR (d/ppm):168 and 166
( C=N), 137-133 (Car), 70 (Csugar).
(Csugar
)
5-(2’-Benzamidophenylene)-1,3,4-triazole-
5-(2’-Benzamidophenylene)-1,3,4-
2-S-glucoside(13b): IR, nmax/ cm-1342O (OH), 3330
(NH), 3067 (CH), 2990 (CHar), 1632 (C=N).1H-NMR,
9.5 (s,1H,NH), 7.8-6.7(m, 9H, Har),5.0(m, 4H,O-H);
3.5 (m,8H, H-C-O), 3.2(d,2H, H2-C-O).13C-NMR
(d/ppm):169 (C=O), 164 ( C=N), 134-120 (Car), 75-
oxadiazole-2-S-glucoside (11b):Yellow syrup.IR,nmax
/
cm-1 342O (OH), 3330 (NH), 3067 (CH), 2990 (CHar),
1
1632 (C=N). H-RMN, 12.3 (s,1H,NH), 8-7.2 (m,
9H, Har),4.0(m, 4H,O-H); 3.7(m,8H, C-H), 3.2(d,2H,
H2-C-O).13C-NMR ,171 (C=O), 167( C=N), 136-
70 (Csugar
)
118(Car), 76-60 (Csugar
)
5-(2’-Aminophenylene)-4-amino-1,3,4-
5-(2’-Aminophenylene)-1,3,4-thiadiazole-
2-S-glucoside (12a): Yellow syrup, . IR, nmax/ cm-1
3360 (broad,OH and NH), 3070 (CH), 2996 (CHar),
1632 (C=N).1H-RMN, 8.6 (s,1H,NH), 7.5-6.5 (m, 4H,
Har),4.2 (m, 4H,O-H);3.57(m,8H, O-C-H), 3.5 (d,2H,
H2C-O).13C-NMR: 175 and 160 ( C=N), 150 (Car), 60
(Csugar).
triazole-2-S-glucoside(14a): recrystallized from
ethanol, m.p. 78°C.IR, nmax/ cm-13423 (OH), 3290
(NH), 3067 (CH), 2998 (CHar), 1630 (C=N).1H-NMR,
9.6 (s, 2H, NH2), 8.2 (s,1H,NH), 8.3-7.6(m, 4H,
Har),4.3(m, 4H,O-H); 3.7(m,8H, H-C-O), 3.2(d,2H,
H2-C-O).13C-NMR (d/ppm): 169 (C=O), 164 ( C=N),
134-120 (Car), 75-70 (Csugar
)
5-(2’-Benzamidophenylene)-1,3,4-
thiadiazole-2-S-glucoside(12b): Yellow syrup,IR,
max/ cm-11 342O (OH), 3330 (NH), 3067 (CH),
2990 (CHar), 1632 (C=N).1H-NMR, 12.3 (s,1H,NH),
8-7.2 (m, 9H, Har),4.0(m, 4H,O-H); 3.7(m,8H, C-H),
3.2(d,2H, H2-C-O).13C-NMR (d/ppm):169 (C=O), 164
5-(2’-Benzamidophenylene)-4-amino-
1,3,4-triazole-2-S-glucoside (14b):IR, nmax/ cm-1 342O
(OH), 3330 (NH), 3067 (CH), 2990 (CHar), 1632
(C=N).1H-NMR, 9.8 (s,1H,NH2),8.9(s,1H,NH), 8-7.2
(m, 9H, Har),4.2(m, 4H,O-H); 3.5 (m,8H, H-C-O),
3.5(d,2H, H2-C-O).13C-NMR (d/ppm):171 (C=O), 163
n
( C=N), 136-118(Car), 78-63 (Csugar
)
( C=N), 134-120 (Car), 72-70 (Csugar)
10(a,b)
9(a,b)
7(a,b)
D-gluc
8(a,b)
D-gluc
D-gluc
D-gluc
N
N
N
N
N
N
N
N
S--
S-gluc
S-gluc
N
R
R
R
N
H
R
S-gluc
O
S
gluc
NH--
11(a,b)
12(a,b)
13(a,b)
14(a,b)
H
H
COPh
Ra =
;
Rb =
N
N
H
Scheme 3: Synthesis of N- and S-glycosides 11(a,b)-14(a,b)

BENHAMMADI et al., Orient. J. Chem., Vol. 32(5), 2567-2576 (2016)
2573
The antibacterial Tests
The filter paper disc method was performed
used.Inhibitory activity was evaluated by measuring
the diameter of clear zone observed around the disc
(in mm).
in duplicate using fresh Mueller Hinton agar medium.
This agar medium was inoculated with 0.5mL of
cultures containing about 106 CFU/mL. Filter paper
discs (5 mm diameter) saturated with solutions of
each compound (concentrations 10µg mL–1 DMSO)
was placed on the indicated agar mediums. The
incubation time was 24 h at 37 °C.The blank test disc
with DMSO and positive reference amykacine were
Minimum inhibition concentration (MIC) Tests
Each 1 mL of the original concentration
[ c ] ( 10 µg mL-1 ) in DMSO of the compounds7(a,b)-
10(a,b) and 12(a,b)were diluted with DMSO for four
times to1/2 [ c ], 1/4[ c ] , 1/8 [ c ] ,1/16[ c ] , and
Table 1: Inhibition of microorganisms by
compounds 7(a,b)-10(a,b)and 12(a,b)in mm
Compounds*
Gram positive
Gram negative
S.aureus
B. cereus
E.coli
P.aeroginosa
7a
7b
8a
8b
9a
9b
0
10
19
21
0
18
11
19
14
0
20
11
20
0
16
0
31
12
0
16
13
17
13
0
11
0
33
19
12
20
21
11
10a
10b
12a
12b
Amykacine
19
25
10
0
22mm
20mm
19mm
0
*concentration 10 µg mL-1
Table 2: minimum inhibition concentrations (MIC)*
Com.
S. aureus
µg mL-1
B. cereus
µg mL-1
E.coli
µg mL-1
P. aeroginosa
µg mL-1
i
ii
iii
iv
i
ii
iii
iv
i
ii
iii
iv
i
ii
iii
iv
7a
7b
8a
8b
9a
9b
10a
10b
12a
12b
-
-
-
-
+
+
-
+
-
+
+
-
-
-
-
+
-
+
-
+
+
-
+
+
+
-
+
-
+
+
+
-
+
-
+
+
+
-
+
-
+
-
+
-
-
-
+
+
-
+
+
+
+
-
+
+
-
+
+
+
+
-
+
-
-
+
+
+
-
-
-
-
+
-
-
-
-
+
-
-
-
-
+
-
+
-
+
-
+
-
+
-
+
+
+
-
+
+
+
+
-
+
+
+
-
+
+
+
+
-
+
+
+
+
+
+
+
+
+
+
+
-
+
+
+
+
+
-
-
+
+
-
+
+
+
+
+
+
-
-
+
-
+
-
i =1/2.C; ii=1/4.C; iii=1/8.C; iv=1/16.C
*(C= initial concentration = 10 µg mL-1)

BENHAMMADI et al., Orient. J. Chem., Vol. 32(5), 2567-2576 (2016)
2574
optical density was measured at 0 Hr, 18 Hr, 24 Hr
and 48 Hr.
and SH bands in the region 3333 cm-1and 2626 cm-1
respectively and for C=S group in 1404cm-1. While
the NMR spectra exhibited signals at 11.2 ppm and
3.3 ppm for the NH and the SH respectively while 13C-
NMR showed both thione at 205ppmandat170.27
ppm for thiol form (N=C-SH) .
RESULTS AND DISCUSSION
Synthesis of hydrazides 3(a,b)
Esters and benzamido acid 2(A-E) and the
corresponding hydrazides 3(a,b) are the common
intermediates for the synthesis described in this work.
They should be available in relatively large quantities
to utilize for further synthesis. In best opportunity,
2-aminomethylbenzoate 2A was obtained by
classical esterification of 2-aminobanzoic acid 1 with
methanol catalysed by H2SO4to give a modest yield
(12%). Since 1 showed a better solubility in ethanol,
classical esterification of 1 using H2SO4 or HCl as
catalysts gave a best yield of 2-aminoethylbenzoate
2B in 30% yield. Generally, the yield of both methyl
and ethyl 2-aminobenzoates 2(A,B) respectively
were considered very poor and unsuitable for
stepwise synthesis of nucleobases 7-10(a,b) (see
Scheme 1). The yield percentages of esterification
been raised firstly by using montmorillonite argil
as catalyst and secondly by protecting the amino
group via N-benzoylation of 1 to give 2-benzamido
benzoic acid 2C prior to esterification. Methyl and
ethyl 2-aminobenzoates 2(A,B) resulted from argil
as catalyst gave higher yield (65%) . Similirlay
2-benzamido methyl and ethylbenzoates 2(D, E)
were also produced in (65%) yield.
Route B: gave rise to the synthesis of
thiadiazolethiols8(a, b)andtriazoles9(a, b).Reaction
of the hydrazides 3(a, b) with ammoniumthiocyanide
resulted into formation of thiosemicarbazides
5(a, b) in excellent yields (86% for 5a and 80% for
5b). IR5a spectra showed a broad band cantered at
3290 cm-1 for NH, 1631 for (C=O) and 1404 for(C=S).
IR5b, 3233 (NH), 1620 (C=O), 1404 (C=S).
Treatment of 5(a,b) with alcoholic KOH
followed by acidification with HCl to pH=1 resulted
into very good yields of thiadiazoles 8a in 80%
yield and 8b in 78% yield. While treatment of
thiosemicarbazides 5(a, b) with aqueous NaOH or
KOH furnished 1,2,4-triazole thiols 9(a,b) in excellent
yields (89% for 9a, and 91% for 9b ).
Route C:dealt with synthesis of 2’-amino-1,
2, 4-triazole thiols 10(a,b) which involving treatment
of the hydrazides 3(a,b) with CS2 in presence of KOH
to give potassium salts of hydrazinic acids 6(a,b).
The latters without further identification when treated
with hydrazine hydrate 64% and KOH resulted the
2’-amino phenelyn-1,2,4-triazole thiols10(a,b) in very
good yields (10a, 93% and 10b, 80%).
Hydrazides 3(a,b) were obtained in
excellent yields 3a( 90%) and 3b (84%) by refluxing
the corresponding esters 2(A-E) with hydrazine
hydrate 51% in methanol solution.Hydrazides 3a and
3b characterized by IR, UV, 1H-NMR and 13C-NMR
(see experimental).
Synthesis of glycosides: 11(a,b)-14(a,b)
The second target of this work was the
synthesis of S-glycosides; those been achieved
by Fischer glycosidation reaction of unprotected
monosaccharides with thiol groups in the presence
of an acid catalyst. Thus, refluxing of nucleobases
7(a,b)-10(a,b) with D- glucose they yielded
S-glucosides 11(a,b)-14(a,b) (see Scheme 3). Thin
layer chromatography have shown formation of
one spot after each reaction with disappearance
of nucleobases spots except for 14(a,b).The latter
always exhibited two very close spots were difficult
to separate by column chromatography.They related
to mixture of S- and N-glucosides. No further work
was done to separate these mixtures. IR and NMR
measurements and melting points supported the tlc
for formation of S-glycosides 11(a,b)-14(a,b).
Synthesis of nucleobases 7(a, b)-10(a, b)
The synthesis of heterocyclic thiols
(nucleobases) 7(a,b)-10(a,b) have been obtained
from the hydrazides3a and 3b into three synthetic
routes A, B and C.
Route A: gave rise to oxadiazoles 4a and
4b, which obtained by treating –indvidually- the
hydrazides 3a and 3b with ethanolic KOH followed by
addition of CS2 under reflux for 14 hours. IR spectra
suggested the presence of keto-enol tautomers
4(a,b)
7(a,b) detected by the presence of NH

2575
BENHAMMADI et al., Orient. J. Chem., Vol. 32(5), 2567-2576 (2016)
benzamido derivative 12b showed no effect toward
Biological Tests
Nucleobases 7(a,b)-10(a,b) and one the two organisms.
type of S-glucosides namely 5-(2’-amino and
2’-benzamidophenylene)-1,3,4-thiadiazole-2-S-
Minimum inhibition concentration (MIC)
glucosides (12a,b) were tested against representative test were performed only on compounds showed an
microorganisms. Two Gram-positive bacteria effect in initial concentration 10 ug.mL-1. Tests were
Staphylococcus aureus ATCC 25923 and Bacillus done in duplicate and the average results shown in
cereus and two Gram-negative bacteria Escherichia Table 2.
coli ATCC 25924, Pseudomonas aeruginosa ATCC
27835.The filter paper disk method (NCCLS) 25was
employed in duplicate for in vitro study of antibacterial
effects and amykacine was used as a positive
CONCLUSIONS
Esterification and benzamidation of 2-amino
reference.The inhibitory effects summarized inTable benzoic acid proved to be synthetically valuable.
1.
2-Benzamido benzoic acid exhibited better solubility
in alcohols which lead to higher yield of esters and
From Table 1, it is shown that almost all ultimately give higher yield of hydrazides 3(a,b).
synthetic nucleobases 7(a,b)-10(a,b) and glycoside Since the hydrazides 3(a,b) were considered the
12a exhibited various effect on the representative key intermediates for the synthesis of nucleobases
bacteria. It seemed that it is difficult to generalize 7(a,b)-10(a,b) and glycosides 11(a,b)-14(a,b). Also
whether there are differences in activity between some differences observed in biological activity
the aminophenylene nucleobases 7a-10a and caused by benzamido derivatives of 2-amino benzoic
their benzamido derivatives 7b-10b. There are only acid particularly in synthesizing nucleobases and
some cases where the effect exceeded the positive S-glucosides 12(a, b).
reference amykacine as in oxadiazole 7a on G(-)
E.coli , activity of thiadiazole 8a upon G(+) B. cereus
and activity of all tested nucleobases 7(a,b)-10(a,b)
on G(-)P. aeraginosa. 5-(2’-Aminophenylene)-1,3,4-
ACKNOWLEDGMENTS
Thanks to Mrs. Karima Abed for typing and
thiadiazole-2-S-glucoside (12a) exhibited a moderate arranging the manuscript.
effect upon G(+) S. aureus and G(-) E. coli ,while its
REFERENCES
1.
2.
Cleaves II H. J. 2011, 1455, Springer Berlin
Heidelberg. doi: 10.1007/978-3-642-11274-
4_1371.
Fiandaca M.J.; Hyldig-Nielsen J.J.; Gildea
B.D.; Coull,J.M. Genome Res. 2001, 11:
609–613.
Hafez H.N.; Hussein H.A.R.;El-Gazzar A.B.A.
Eur. J. Med. Chem. 2010, 45(9): 4026–4034.
Mohamed A.M.;Al-Qalawi H.R.M.;El-Sayed
W.E.; Arafa W. A. A. Acta Poloniae Pharm.
Drug Res. 2015, 72: 307-331.
Khalil N. S.A.M. Carbohydrate. Res. 2006,
341(13): 2187–2199.
Aderiye B.I.; Oluwole O. A. Agri. Biol. Sci. J.
2015, 1(5): 206-216. http://www.aiscience.
org/journal/absj.
Pharm. Bull. 2016, 64(5): 439-450. doi:
org/10.1248/cpb.c15-01005.
Sharonova T.; Baykov S.; Dorogov M.
IRJPAC. 2016, 10(3): 1-5. doi: 10.9734/
IRJPAC/2016/22429.
Shigeta S.;Mori S.;Baba M.;Ito M.;Honzumi
k.; Nakamura K.; OshitaniYNumazaki
H.; Matsuda A and Obara T. Antimicrob.
Agents Chemother. 1992, 36(2): 435-439.
doi: 10.1128/AAC.36.2.435.
Krajczyk A.; Kulinska K.; Kulinski T.; Hurst
B.L.; Day C. W.; Smee D.F.; Ostrowski T.;
Januszczyk P.; Zeidler J. Antiviral Chem.
Chemotherapy 2014, 23:161-171. doi:
10.3851/IMP256.
8.
9.
3.
4.
10.
11.
5.
6.
Kini GD.; Robins RK.; Avery T.L. J.Med.
Chem. 1989, 32 (7):1447–1449.doi:10.1021/
7.
Azab ME.; Rizk S.A.; Mahmoud N. F. Chem.

BENHAMMADI et al., Orient. J. Chem., Vol. 32(5), 2567-2576 (2016)
2576
jm00127a008.
18.
19.
20.
Amarouche L.; Taieb Brahimi F.; Othman A.
A. J.Chem.Pharm.Res.2016, 8(4):896-900.
ISSN: 0975-7384.
Othman AA.;Kihel M.;Amara S.Arab J Chem.
2014, doi.org/10.1016/j.arabjc.2014.09.003
(references theirin).
S. M.,Vaughan M. D., Withers S. G., Current
Opinion in Chemical Biology. 2006, 10(5):
509–519. doi:10.1016/j.cbpa.2006.07.015.
Kumar S.; Singh D.; Dutta M. J. Oilseed
Brassica. 2014, 5(2): 87-95.
JohnsonS.;TanakaF.Org.Biomol.Chem.2016,
14: 259-264. doi: 10.1039/ C5OB02094H.
Zavareh E. R.; Hedayati M.; Rad L. H.;
Shahhosseini S.; Faizi M.; Tabatabai SA.
Iran J Pharm Res. 2014, 13(Suppl) : 51-9.
Chavhan NM.; Badadhe PV.; Shelke SN . Int
J Innov Res Sci Eng Technol. 2015, 4: 417-
421. doi: 10.15680 /IJIRSET.2015.0402027.
Dallal MMS, Doyle MP, Rezadehbashi M,
Dabiri H, Sanaei M, Modarresi S, Bakhtiari R,
Sharifiy K, Taremi M, Zali MR, Sharifi-Yazdi
MK : Food Control . 2010, 21: 388–392. doi:
10.1016/j.foodcont.2009.06.001
12.
13.
14.
Witkoviski JT.; Robins RK. J. Org. Chem.
1970, 35(8): 2635-2641. doi: 10.1021/
jo00833a033.
Cipolla L. ; Palma A.; La Ferla B.;Nicotra F.
J. Chem. Soc., Perkin Trans. 2002, 1. 2161-
2165. doi: 10.1039/B206623.
Brito-Arias M.“SynthesisandCharacterization
of Glycosides”. Springer international
publishing, Swistzerland,. 2016, 81-168. doi
10.1007/978-3-319-32310-7_2.
Schmidt R. R.; Angew. Chem. International.
1986, 25(3): 212–235, doi: 10.1002/
anie.198602121.
Nicolaou K. C. ; Chucholowski A.; Dolle
R. E.; Randall J. L. J. Chem. Soc., Chem.
Commun. 1984, 1155-1156. doi: 10.1039/
C39840001155.
Almasirad A.; Tabatabai SA.; Faizi
M.; Kebriaeezadeh A.; Mehrabi N.; Dalvandi
A.; Shafiee, Bioorg.Med. Chem.Lett. 2004,
14(24): 6057-6059. doi.org/10.1016/j.
bmcl.2004.09.072 : 6057.
21.
22.
23.
15.
16.
24.
25.
17.