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Synthesis and Antimicrobial Activity of Novel α- Aminophosphonates Bearing Pyrazoloquinoxaline Moiety

Authors:
  • Aswan University- faculty of veterinary medicine

Abstract and Figures

Syntheses of novel N-protected α-aminophosphonates 6 were achieved with high yields through lithium per chlorate catalyzed one-pot three component reaction process. It involves the reaction of aryl substituted Quinoxalinealdehydes, benzyl carbamate, aniline, p-methoxy aniline, p-chloro aniline, p-methyl aniline, propyl amine, pentyl amine, p-amino benzoic acid, 1,4-phenylenediamine, amino uracil, N-glycosyl amine and triphenylphosphite using lithium perchlorate as Lewis acid catalyst in dry dichloromethane at room temperature. A mechanism for this condensation reaction is proposed. Cleavage of the N-phenyloxycarbonyl group under acid hydrolysis afforded the free α-aminophosphonates 8 in quantitative yields. The structures of all new compounds were established by IR, ¹HNMR, 13 CNMR and mass spectral data. All the synthesized compounds were screened for in vitro antibacterial activity and most of them showed potency against both gram positive and gram negative bacteria.
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Int. J. Pharm. Sci. Rev. Res., 34(1), September – October 2015; Article No. 33, Pages: 205-213 ISSN 0976 – 044X
International Journal of Pharmaceutical Sciences Review and Research
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Mohamed A. Hamed 2, Ahmed A. El Gokha1, Alaa Fathy A. Ahmed1, Mohamed Sabry Abd Elraheam Elsayed3, R. Tarabee3,
Ahmed El S. Abdel Megeed1, Ibrahim El-Tantawy El Sayed1
1Chemistry Department, Faculty of Science, El-Menoufeia University, Shebin El-Kom, Egypt.
2Chemistry Department, Faculty of Science, Tanta University, Tanta, Egypt.
3Department of Bacteriology, Mycology and Immunology Faculty of Veterinary Medicine University of Sadat City, Menoufia, Egypt.
*Corresponding author’s E-mail: ibrahimtantawy@yahoo.co.uk
Accepted on: 21-07-2015; Finalized on: 31-08-2015.
ABSTRACT
Syntheses of novel N-protected α-aminophosphonates 6 were achieved with high yields through lithium per chlorate catalyzed one-
pot three component reaction process. It involves the reaction of aryl substituted Quinoxalinealdehydes, benzyl carbamate, aniline,
p-methoxy aniline, p-chloro aniline, p-methyl aniline, propyl amine, pentyl amine, p-amino benzoic acid, 1,4-phenylenediamine,
amino uracil, N-glycosyl amine and triphenylphosphite using lithium perchlorate as Lewis acid catalyst in dry dichloromethane at
room temperature. A mechanism for this condensation reaction is proposed. Cleavage of the N-phenyloxycarbonyl group under acid
hydrolysis afforded the free α-aminophosphonates 8 in quantitative yields. The structures of all new compounds were established
by IR, ¹HNMR, 13CNMR and mass spectral data. All the synthesized compounds were screened for in vitro antibacterial activity and
most of them showed potency against both gram positive and gram negative bacteria.
Keywords: Pyrazoloquinoxaline, amines, triphenylphosphite, Lewis Acid, α-aminophosphonates, Antimicrobial Activity.
INTRODUCTION
rganophosphorus compounds have found a wide
range of applications in the areas of industrial,
agricultural, and medicinal chemistry owing to
their biological and physical properties as well as their
utility as synthetic intermediates1. α-Functionalized
phosphonic acids are valuable intermediates for the
preparation of medicinal compounds and synthetic
intermediates2–4. Among α-functional phosphonic acids,
α-aminophosphonic acids are an important class of
compounds that exhibit a variety of interesting and useful
properties. α-Aminophosphonic acids I, as structural
mimics of α-amino acids II (Fig.1), exhibit a broad
spectrum of biological activities5-12.
These compounds have already been found to act as
antibacterial agents, neuroactive compounds, anticancer
drugs, and pesticides, with some of them already
commercialized13-18. In this context, the therapeutic
potential for modified α-aminophosphonates with
improved pharmacokinetic properties, potency or
spectrum, and lower side effects, prompted us to start a
synthetic program to explore new Quinoxaline α -
aminophosphonate conjugates. We focused on
Quinoxaline and its derivatives because it is an important
class of compounds and attracted widespread attention
due to their pharmacological properties, being reported
to have a large spectrum of biological effects, especially
analgesic, antibacterial, antifungal, anticancer and anti-
inflammatory properties. In this paper we would like to
present the synthesis of novel Quinoxaline modified α-
aminophosphonates conjugates and biological evaluate
as antibacterial activity.
MATERIALS AND METHODS
All 1HNMR and 13CNMR experiments (solvent DMSO)
were carried out with a 300 MHz at the University of Ulm,
Germany, Okayama university, Japan. Chemical shifts are
reported in part per million (ppm) relative to the
respective solvent or tetramethylsilane (TMS). The mass
spectroscopy experiments and IR spectroscopy were
performed at Cairo University, Egypt. Melting points were
recorded on Stuart scientific melting point apparatus and
are uncorrected. The microanalysis was performed in
department of Bacteriology, Mycology and Immunology
Faculty of Veterinary Medicine University of Sadat City,
Menoufia, Egypt. All reactions were followed by thin layer
chromatography (TLC) on kiesel gel F254 precoated plates
(Merck). Starting materials, MeOH, DMF, acetonitrile,
CH2Cl2, hexane and diethyl ether were either
commercially available as reported in literature.
Synthesis of (1S, 2S)-1-(1-phenyl-1H-pyrazolo [3, 4-b]
quinoxalin-3-yl) propane-1, 2, 3-triol: (.i)
A solution of L-glucose (1.8 gm., 0.01 mol.) in water (800
ml.) was heated with O-phenylenediamine (1.08 gm., 0.01
mol.), phenyl hydrazine hydrochloride (7.2 gm., 0.01
mol.), glacial acetic acid (11 ml.) and 0.2 gm. Sodium
acetate. The reaction mixture was treated as previously
mentioned method; Recrystallization from ethanol gave
Synthesis and Antimicrobial Activity of Novel α- Aminophosphonates
Bearing Pyrazoloquinoxaline Moiety
O
Research
Article
Int. J. Pharm. Sci. Rev. Res., 34(1), September – October 2015; Article No. 33, Pages: 205-213 ISSN 0976 – 044X
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yellow needles and Show the following data; MP. =235 °C
Yield= 80%, Infra-red spectra of compound (.i) show:
3461.6, 2932.2, 1598.7, 1500.35 and 757.88 cm-1, 1H NMR
(DMSO-400 MHz): δ ppm = 3.43-3.54(m, 2H , 3-H), 4.34-
4.45 (m, 1H , 2-H), 4.64 (t, 1H , J= 5.49 Hz, OH), 4.88 (d.1H
,J= 5.56 Hz, OH),5.34 (t.1H ,J= 5.58 Hz, OH),5.56 (d.1H ,J=
6.18 Hz, 1H),7.35-8.46 (m, 9H , H aroma.), 13CNMR
(DMSO-d): δ ppm = 60.77-95.89 (Sugar-C), 119.52-144.90
(aroma.-C).
Synthesis of 1-phenyl-1H-pyrazolo [3, 4-b] quinoxaline-
3-carbaldehyde: (.ii)
A solution of periodic acid (2.28 gm. 1mol.) in water (100
ml.) was added drop wise during 3 h. at room
temperature to a stirred suspension of (.i) (0.336 gm.
0.001 mol.) in water (50 ml.).
The mixture is then stirred for 20 h. the yellow precipitate
was filtered. Off and successively washed with water and
50 % propanol, dried, recrystallization from 80% propanol
gave yellow crystals and Show the following data Mp. 150
°C; Yield: (85%), Infra-red spectra of compound (.ii) show:
1701.27, 1598.7, 1500.13, and 761.35 cm-1, 1HNMR
(DMSO, 400 MHz) δ = 7.26– 8.49 (m, 9H, H aroma), 10.49
(s, 1H, formyl 1-H), 13CNMR (DMSO-d): δ ppm = 119.52-
144.90 (aroma.-C), 185.19 (1 CHO).
The mass spectra show the molecular ion peak at m/e =
274 [M] + (84.47 % is base peak), The ion peak at m/e =
245 (- CHO) [M] +.
Reaction of 1-phenyl-1H-pyrazolo [3, 4-b] quinoxaline-3-
carbaldehyde with Amines and Triphenylphosphite;
General Procedure:
1-phenyl-1H-pyrazolo [3, 4-b] quinoxaline-3-carbaldehyde
(1mlmol.), amines (1mlmol.) and triphenylphosphite
(1mlmol.) Were dissolved in (5 ml) of dry
dichloromethane. The Lewis acid* (10 mol. %) is added in
one portion.
The mixture was stirred at room temperature, until TLC
analysis showed the complete consumption of 1-phenyl-
1H-pyrazolo 3,4-b quinoxaline-3-carbaldehyde after 72 h.
The dichloromethane is then evaporated and the residue
dissolved in diethyl ether (10 ml) the product was
precipitated from this solution by storing at – 20 C for 3 –
6 h. followed by the collection of the precipitate by
filtration afford the protected aminophosphonates 5 in
good to excellent yields.
* LiClO4 was added as 1 mol. Solution in dry
dichloromethane.
Di phenyl ((1-phenyl -1H- pyrazolo [3, 4-b] qunaxolin-3-
yl) (p-tolylamino) methyl) phosphonate: (.iii)
Show the following data MP. = 98-102 °C, Yield = 85 %,
Infra-red spectra of compound (.iii) show: 3439.42,
2863.52, 1643.87, 1511.07, 1151.16, 1118.46 and 761.35
cm-1, 1HNMR (DMSO, 400 MHz) δ =5.48-5.62 (s, 1H, NH),
3.1 (s, 3H, CH3), 4.5-4.7 (m, 1H, CHP), 7.36-7.65(m, 10H, H
aroma.), 7.88-8.41(m, 5H, H aroma.), The mass spectra
show the molecular ion peak at m/e = (597[M] +, 5.54%),
the base ion peak at m/e = 215[M] + - (C23H18N53.91%).
Di phenyl ((((4-methoxy phenyl) amino (1-phnyl – 1H-
pyrazolo [3, 4, b] quinaxolin-3-yl) methyl) phosphonate:
(.iv)
Show the following data MP. = 100 °C, Yield = 87%, Infra-
red spectra of compound (.iv) show: 3442.89, 2851.26,
1643.57, 1506.71, 1142.99, 1116.52 and 758.83 cm-1,
1HNMR (DMSO, 400 MHz) δ =2.5-2.8 (s,3H,CH3O),5.12-
5.15 (m, 1H,CHP), 5.7-6.0 (s,1H,NH) ,7.36-7.65(m, 10H,H
aroma.),7.88-8.44(m, 5H,H aroma.), The mass spectra
show the molecular ion peak at m/e = (614[M] +, 77.59%),
the base ion peak at m/e = 370[M] +-(C15H9N465.52%).
Diphenyl (((4-chlorophenyl) amino) (1-phenyl-1H-
pyrazolo [3, 4-b] quinoxalin-3-yl) methyl) phosphonate:
(.v)
Show the following data MP. = 98-100 °C, Yield = 80 %,
Infra-red spectra of compound (.v) show: 3438.82,
1642.41, 1503.9, 1147.42, 1114.31 and 758.46 cm-1,
1HNMR (DMSO, 400 MHz) δ =5.07-5.14 (m, 1H, CHP),
6.25-6.32 (s, 1H, NH), 7.37-7.64(m, 10H, H aroma.), 7.89-
8.42(m, 5H, H aroma.), 13CNMR (DMSO-d): δ ppm =
119.62-148.68 (C- aroma.), 153.26-159 (1 NH), The mass
spectra show the molecular ion peak at m/e = 618[M] +,
84.07%), the base ion peak at m/e = 373[M] + -
(C15H9N469.03%).
Diphenyl ((1-phenyl-1H-pyrazolo [3, 4-b] quinoxalin-3-yl)
(phenylamino) methyl) phosphonate: (.vi)
Show the following data MP. = 95-98 Yield = 80%, Infra-
red spectra of compound (.vi) show: 3442.89, 1643.57,
1510.79, 1151.16, 1090.46 and 767 cm-1, 1HNMR (DMSO,
400 MHz) δ =5.06-5.92 (m, 1H, CHP), 6.69-6.71 (s, 1H,
NH), 7.38-7.65(m, 10H, H aroma.), 7.90-8.43(m, 5H, H
aroma.), The mass spectra show the molecular ion peak
at m/e = 583.58[M] +, 45.08%), the base ion peak at m/e =
338[M] + - (C15H8N473.77%).
Diphenyl ((1-phenyl-1H-pyrazolo [3, 4-b] quinoxalin-3-yl)
(propylamino) methyl) phosphonate: (.vii)
Show the following data MP. = 198 – 201 °C, Yield = 75%,
Infra-red spectra of compound (.vii) show: 3438.21,
2984.51, 1638.28, 1503.45, 1152.52, 1006.55 and 763 cm-
1, 1HNMR (DMSO, 400 MHz) δ =1.98-2.0 (m,4H,CH2), 3.70-
3.80 (m,3H,CH3), 5.40-5.50 (m, 1H,CHP), 5.80-6.0
(s,1H,NH) ,7.13-7.66(m, 10H,H aroma.),7.66-8.44(m, 5H,H
aroma.), The mass spectra show the molecular ion peak
at m/e = 549.56[M] +, 15.61%), the base ion peak at m/e =
215[M] +–(C19H18N510.87 %).
Diphenyl ((pentylamino) (1-phenyl-1H-pyrazolo [3, 4-b]
quinoxalin-3-yl) methyl) phosphonate: (.viii)
Show the following data MP. = 230-233°C, Yield = 75 %,
Infra-red spectra of compound (.viii) show: 3437,
1636.93, 1502.62, 1147.07, 1009.28 and 762.92 cm-1,
1HNMR (DMSO, 400 MHz) δ =1.98-2.0 (m,8H,CH2), 3.50-
Int. J. Pharm. Sci. Rev. Res., 34(1), September – October 2015; Article No. 33, Pages: 205-213 ISSN 0976 – 044X
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3.60 (m,3H,CH3), 5.06-5.08 (m, 1H,CHP), 5.80-6.0
(s,1H,NH) ,7.23-7.91(m, 10H,H aroma.),7.91-8.44(m, 5H,H
aroma.), The mass spectra show the molecular ion peak
at m/e = 577.61[M]+, 71.82% ), the base ion peak at m/e =
332[M]+–(C15H8N451.82 %).
Diphenyl((1-phenyl-1H-pyrazolo[3,4-b]quinoxalin-3-
yl)(((2R,3R,4R,5R,6R)-2,4,5-trihy droxy-6-
(hydroxymethyl)tetrahydro-2H-pyran-3-yl)amino)
methyl) phosphonate: (.ix)
Show the following data MP = 155 C, Yield = 65 %, Infra-
red Spectra of compound (.ix) show: 3422, 1627.63,
1498.42, 1145.51, 1110.8 and 756.923 cm-1, 1HNMR
(DMSO, 400 MHz) δ =4.47 (d,2H,4-OH), 5.07-5.09 (m,
1H,CHP), 6.90-7.0 (s,1H,NH) ,7.37-7.64(m, 10H,H
aroma.),7.99-8.42(m, 5H,H aroma.).
The mass spectra show the molecular ion peak at m/e =
669.62[M] +, 25.49%), the base ion peak at m/e = 178[M]
+–(C28H19N4O3P 23.20 %).
4-(((diphenoxyphosphoryl) (1-phenyl-1H-pyrazolo [3, 4-
b] quinoxalin-3-yl) methyl) amino) benzoic acid: (.x)
Show the following data MP. = 130C, Yield = 85 %, Infra-
red spectra of compound (.x) show: 3427, 1602.56,
1498.42, 1144.55, 1089.58 and 756.923 cm-1, 1HNMR
(DMSO, 400 MHz) δ = 5.06-5.08 (m, 1H,CHP), 5.84 (s,
1H,H-acid),6.52-6.55 (s,1H,NH) ,7.38-7.66(m, 10H,H
aroma.),7.67-8.44(m, 5H,H aroma.), 13CNMR (DMSO-d): δ
ppm = 112.61-146.31 (C- aroma.), 167.73-178.40 (1-COO).
The mass spectra show the molecular ion peak at m/e =
627.58[M] +, 17.60%), the base ion peak at m/e =
385[M+2] +–(C15H8N423.46 %).
Diphenyl(((2,6-dioxo-1,2,3,6-tetrahydropyrimidin-4-
yl)amino)(1-phenyl-1H-pyrazolo[3,4-b]quinoxalin-3-
yl)methyl)phosphonate: (.xi)
Show the following data MP. = 167-170C Yield = 60%,
Infra-red spectra of compound (.xi) show: 3415, 1710.55,
1627.63, 1501.31, 1238.08, 896.74 and 759.82 cm-1,
1HNMR (DMSO, 400 MHz) δ = 5.08-5.10 (m, 1H,CHP), 6.27
(s,1H,NH) ,7.37-7.66(m, 10H,H aroma.),7.66-8.43(m, 5H,H
aroma.),10.21(s, 1H,OH),11.12(s, 1H,NH), 13CNMR
(DMSO-d): δ ppm = 119.52-148.85 (aroma. - C), 155.43-
167.73 (2 C=O).
Tetraphenyl((1,4-phenylenebis(azanediyl))bis((1-phenyl-
1H-(pyrazolo[3,4-b]quinoxalin-3-yl)
methylene))bis(phosphonate): (.xii)
Show the following data MP. = 145-148C. Yield = 80 %,
Infra-red spectra of compound (.xii) show: 3428.81,
1630.52, 1499.38, 1230.36, 902.523 cm-1 and 757.888 cm-
1, 1HNMR (DMSO, 400 MHz) δ = (m, 1H,CHP) 4.47-
4.49,5.07-5.10(m, 1H,CHP), 5.87(s,1H,NH) ,7.37-7.66(m,
20H,H aroma.),7.88-8.44(m, 8H,H aroma.).
The mass spectra show the molecular ion peak at m/e =
1089.04[M] +, 66.23%), the base ion peak at m/e =
843[M+2] +–(C15H8N429.80%).
Diphenyl (((4-aminophenyl) amino) (1-phenyl-1H-
pyrazolo [3, 4-b] quinoxalin-3-yl) methyl) phosphonate:
(.xiii)
Show the following data MP. = 200-203C. Yield = 90%,
Infra-red spectra of compound (.xiii) show: 3457,
1624.93, 1501.31, 1235.18, 900.594 and 756.923 cm-1,
1HNMR (DMSO, 400 MHz) δ = 5.07-5.09 (s, 2H, NH2), 5.87
(m, 1H, CHP), 6.70 (s, 1H, NH), 7.38-7.92(m, 10H, H
aroma.), 8.00-8.44(m, 5H, H aroma.), 13CNMR (DMSO-d):
δ ppm = 119.40-148.82 (C- aroma.), The mass spectra
show the molecular ion peak at m/e = 577.61[M] +,
71.82%), the base ion peak at m/e = 354[M] +–(C15H8N4
41.95 %).
Benzyl((diphenoxyphosphoryl)(1-phenyl-1H-
pyrazolo[3,4-b]quinoxalin-3-yl)methyl) carbamate: (.xiv)
Show the following data MP. = 125C Yield = 65%, Infra-
red spectra of compound (.xiv) show: 3074.44, 1702.66,
1644.06 , 1500.74, 1194.86, 1037.27 and 760.89 cm-1,
1HNMR (DMSO, 400 MHz) δ = 4.98 (m, 1H, CHP), 5.14-
5.22 (m, 2H, phCH2O), 6.12 (s, 1H, NH), 7.17-7.65(m, 10H,
H aroma.), 13CNMR (DMSO-d): δ ppm = 119.47-149.73
(aroma. - C), 155.98-180.00 (1 C=O).
Diphenyl (amino(1-phenyl-1H-pyrazolo[3,4-b]quinoxalin-
3-yl)methyl)phosphonate: (.xv)
A solution of ((diphenoxyphosphoryl)(1-phenyl-1H-
pyrazolo[3,4-b]quinoxalin-3-yl)methyl) carbamate ( 0.128
g – 0.199mlmol.), 3-4 drop of tri ethyl amine and 2-4 drop
of (HBr/Acetic) in (5 ml) of dry dichloromethane .The
mixture was stirred at room temperature, until TLC
analysis showed the complete consumption after 1-2 hr.
the dichloromethane is then evaporated. Show the
following data MP. = 145C yield 65%, Infra-red spectra of
compound (.xv) show: 3413, 1594.84, 1495.53, 1122.37,
1031.73 and 757.888 cm-1, 1HNMR (DMSO, 400 MHz) δ =
5.01-5.14 (m, 1H, CHP), 5.17-5.21 (m, 2H, NH2), 7.33-7.66
(m, 10H, H aroma.), 7.95-8.43(m, 5H, H aroma.).
Synthesis of diphenyl ((1-phenyl-1H-pyrazolo [3, 4-b]
quinoxalin-3-yl) ((4-(3-phenylthioureido) phenyl) amino)
methyl) phosphonate: (.xvi)
A solution of Diphenyl (((4-aminophenyl) amino) (1-
phenyl-1H-pyrazolo [3, 4-b] quinoxalin-3-yl) methyl)
phosphonate (0.334mlmol) and isothiocyanatobenzene
(0.334mlmol) were dissolved in (5 ml) of dry
dichloromethane. The mixture was stirred at room
temperature, until TLC analysis showed the complete
consumption after 2-4 hr. the dichloromethane is then
evaporated and the residue dissolved in di ethyl ether (10
ml) the product was precipitated from this solution by
storing at – 20 C for 3 – 6hr. Show the following data MP.
= 187 190 C Yield = 80 %, Infra-red spectra of
compound (.xvi) show: 3460, 3228.25, 1709.59, 1597.73,
1502.28, 1237.11, 906.379 and 756.923 cm-1, 1HNMR
(DMSO, 400 MHz) δ = 3.68-3.88 (d,2H,NH) ,4.47-4.64 (m,
1H,CHP), 5.07-5.08 (s,1H,NH) ,5.85 (s,1H,SH) ,7.33-
7.68(m, 10H,H aroma.),7.89-8.45(m, 8H,H aroma.), The
Int. J. Pharm. Sci. Rev. Res., 34(1), September – October 2015; Article No. 33, Pages: 205-213 ISSN 0976 – 044X
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mass spectra show the molecular ion peak at m/e =
733.78[M] +, 20.44%), the base ion peak at m/e =
489[M+1] +–(C15H8N418.87 %).
2-(1, 3-dioxoisoindolin-2-yl) propanoic acid: (.xvii)
A mixture of phthalic anhydride (10mlmol.) and L-alanine
(10mlmol.) mixed to completely fusion in sand passes.
Then leave to cold and boiling the product with ethanol.
Filtered as a hot, take the filtrate and cold it Show the
following data MP. =153 °C. Yield = 90 %, Infra-red
spectra of compound (.xvii) show: 3208 cm-1, 1699.94,
1531.2 cm-1 and 726.07 cm-1, 1HNMR (DMSO, 400 MHz) δ
= 1.03-1.06 (m, 2H, CH2), 3.41-3.46 (m, 3H, CH3), 7.79-
7.88(m, 5H, H aroma.), 12.38(s, 1H, H acid), 13CNMR
(DMSO-d): δ ppm = 123.06-167.66 (aroma. - C), 172.19 (1
C=O).
Diphenyl((2-(1,3-dioxoisoindolin-2-yl) propanamido)(1-
phenyl-1H-pyrazolo[3,4-b] quinoxalin-3-yl)
methyl)phosphonate: (.xviii)
A solution of diphenyl (amino(1-phenyl-1H-
benzo[g]pyrazolo[3,4-b]quinoxalin-3-yl)methyl)
phosphonate (0.179 mlmol.), 2-(1,3-dioxoisoindolin-2-yl)
propanoic acid (0.456mlmol.) and 2 drop of triethylamine
in (5 ml) of dry dichloromethanein presence of little
amount of TBTU as a catalyst . The mixture was stirred at
room temperature, until TLC analysis showed the
complete consumption. The dry dichloromethane is then
evaporated and the residue dissolved in (10 ml) of di
ethyl ether the product was precipitated from this
solution by storing at 20 C for 3 – 6h. Show the
following data MP. = 140 C Yield = 65 %, Infra-red spectra
of compound (.xviii) show: 2955.3, 1733.29, 1599.02,
1502.62, 1186.76, 1003.02 cm-1 and 760.38 cm-1, 1HNMR
(DMSO, 400 MHz) δ = 2.49-2.60(q, 1H,CH),3.09-3.10(d,
3H,CH3), 5.10 (m, 1H,CHP), 6.60 (s,1H,NH) ,7.10-7.20(m,
10H,H aroma.),7.25-7.84(m, 8H,H aroma.).
Diphenyl ((2-aminopropanamido) (1-phenyl-1H-pyrazolo
[3, 4-b] quinoxalin-3-yl) methyl) phosphonate: (.xix)
To a solution of biphenyl ((2-(1, 3-dioxoisoindolin-2-yl)
propanamido) (1-phenyl-1H-pyrazolo [3, 4-b] quinoxalin-
3-yl) methyl) phosphonate (0.299mlmol.) and hydrazine
hydrochloride (0.8mlmol.) add mixture of tri ethyl amine
(0.7 ml) and dry methanol (5ml) drop by drop to the
previous solution. At reflux in water bass at 40 C with
stirring at 4h. Leave it to cold. Add (30ml) of water and
(30ml) of ethyl acetate. The lower layer formed with
(water, tri ethyl amine and methanol). The upper layer
formed with (ethyl acetate and the product). Using
separating funnel to separation of the tow layer. By using
rotary vapor to the upper layer to the dispersion of ethyl
acetate, and by using di ethyl ether formed the product.
Show the following data MP. = 135 C Yield = 65 %, Infra-
red spectra of compound (.xix) show: 3437, 1636.93,
1502.62, 1147.07, 1114.38 and 762.92 cm-1, 1HNMR
(DMSO, 400 MHz) δ =1.28(d, 3H,CH3),3.74(q, 1H,CH), 4.90
(m, 1H,CHP), 5.11 (d,2H,NH2) ,7.21-8.24(m, 19H,H
aroma.),8.03(d, 1H,NH).
Diphenyl ((2-benzamidopropanamido) (1-phenyl-1H-
pyrazolo [3, 4-b] quinoxalin-3-yl) methyl) phosphonate:
(.xx)
To a solution of diphenyl ((2-aminopropanamido) (1-
phenyl-1H-pyrazolo [3, 4-b] quinoxalin-3-yl) methyl)
phosphonate (0.099-mmol), benzoyl chloride
(0.099mlmol.) And 2 drop of tri ethyl amine in (5 ml) of
dry dichloromethane the mixture was stirred at room
temperature, until TLC analysis showed the complete
consumption after 3-4 hr. Then CH2Cl2 was evaporated
and the residue dissolved in di ethyl ether (10 ml) the
product was precipitated from this solution by storing at
20 C for 3 – 6hr. Show the following data MP. = 186-
170 C Yield = 65 %, Infra-red spectra of compound (.xx)
show: 3437, 1636.93, 1502.62, 1147.07, 1114.38 and
762.92 cm-1, 1HNMR (DMSO, 400 MHz) δ = 1.48(d,
3H,CH3),4.71 (q, 1H,CH), 4.90 (m, 1H,CHP), 5.11
(d,2H,NH2) ,7.21-8.03(m, 24H,H aroma),8.03,8.45(d,
2H,NH), The mass spectra show the molecular ion peak at
m/e = 682.66[M]+, 34.84% ), the base ion peak at m/e =
437[M]+–(C15H8N4 17.74 %).
Int. J. Pharm. Sci. Rev. Res., 34(1), September – October 2015; Article No. 33, Pages: 205-213 ISSN 0976 – 044X
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RESULTS AND DISCUSSION
The synthesis of mono- and disubstituted diphenyl α-
aminophosphonates 5 were accomplished in good yield
using 1-phenyl-1H-pyrazolo3,4-b quinoxaline-3-
carbaldehyde, a different and triphenylphosphite in the
presence of a Lewis acid such as lithium per chlorate
according to scheme 2. The required aldehydes needed
for this study were synthesized according to published
method19 using Vilsmeier reagent as shown in scheme 1.
Having a diverse series of quinoxalinealdehyde
derivatives affording the opportunity to obtain a various
structures diversity of α-amino- phosphonates 5 by a fast
and convenient one-pot three component reaction route
according to scheme 2.
Optimal conditions for the Lewis acid were found to be 10
mol% in dichloromethane. At 5 mol%, the reaction
afforded the same yield but required longer reaction
times. The reactions are clean and complete within hours.
The reaction conditions are very mild and α-
aminophosphonates are exclusively formed without the
formation of any undesired side products another
important feature of this reaction is the survival of a
variety of functional groups such as ester under the
reaction conditions.
Moreover, the mechanism of this reaction has not been
investigated in detail. We suppose that after reaction of
the carbonyl compound with the amines in presence of
Lewis acid catalyst,
The acylimine intermediate III is attacked by
nucleophilicphosphite with the formation of a
phosphonium intermediate IV and that both reactions are
catalyzed by the Lewis acid. Reaction of phosphonium
intermediate IV with water affords the target compound
5 after elimination of phenol as shown in scheme 3,4.
Finally, in especially reaction with benzyl carbamated
protection or cleavage of the phenyloxycarbonly group by
acidic hydrolysis using HBr/acetic cleanly affords the free
α-aminophosphonates 7 in high yields as shown in
scheme 5. In all cases, the reaction proceeded smoothly
at ambient temperatures with high selectivity.
In summary, we found that a Lewis acid such as LiClO4
effectively promoted the condensation of heterocyclic
aldehydes bearing quinoxaline moiety with benzyl
carbamate and triphenylphosphite or at room
temperature. In addition to we have demonstrated a
novel and efficient protocol for the synthesis of -
aminophosphonates which can serve as peptide mimetic.
The method is effective for heterocyclic aldehydes such
as quinoxalinealdehyde and provides excellent yields of
Int. J. Pharm. Sci. Rev. Res., 34(1), September – October 2015; Article No. 33, Pages: 205-213 ISSN 0976 – 044X
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210
the products, which makes it useful and attractive
process for the synthesis of α-aminophosphonates. It is
believed that this method presents a better and more
practical alternative to the existing methodologies20 for
the synthesis of -aminophosphonates.
Antimicrobial Screening
Antibacterial agents are crucial in reducing the widely
spreaded global burden of infectious bacterial diseases.
As consequence to development of resistant pathogens
and spread, the efficacy of many antibiotics is diminished.
This type of bacterial resistance to the antimicrobial
agents possesses critical threat to public health, and for
all kinds of antibiotics, including the major last-resort
drugs, the frequencies of resistance are increasing
worldwide (Mandal and Mandal, 2011) and Mandal
(2009). Pseudomonas aeruginosa is widely spreaded an
opportunistic Gram negative pathogen Implicated in
multiple infections as respiratory, urinary, gastrointestinal
infections, keratitis, otitis media, and bacteremia in
patients with compromised host defenses [e.g., burn,
cancer, cystic fibrosis (CF) and HIV]. Significant morbidity
and mortality are the main coincidence of such notorious
infectious agent Morita (2014).
Staphylococcus aureus affects humans is a major cause
of morbidity, and economic loss in production in
companion and food animals causing community
acquired and nosocomial infections Rubin (2011).
Wide spectrum of diseases in man and animals are
caused by Aeromonas species Ghenghesh. Recently,
some motile Aeromonas species are becoming food and
waterborne pathogens of great importance Ansari et al.,
(2011). They have been associated with several food-
borne outbreaks and are progressively being isolated
from patients with traveler’s diarrhea Von Graevenitz et
al., (2007).
Pasteurella multocida It is azoonotic Gram negative
bacterium responsible for arrange of infections in
domestic animals, fowl cholera in domestic and wild
birds, bronchopneumonia and hemorrhagic septicemia in
bovids, atrophic rhinitis in porcines and snuffles in rabbits
causing substantial economic losses (Steen) and (Hunt).
Vibrio cholerae O1 and enterotoxigenic Escherichia coli
(ETEC) considered two major bacterial pathogens
responsible for a high proportion of diarrhoeal disease
and death in adults and children in many countries in
Africa and Asia (Svennerholm, 2011) also, Shiga toxin–
producing Escherichia coli (STEC) are a leading cause of
bacterial enteric infections Brooks.
Salmonella enterica species are widely dispersed in
nature and are common inhabitants and highly adapted
to the intestinal tract of domesticated and wild mammals,
reptiles, birds, and even insects. S. enterica Typhi causes
typhoid fever only in humans, whereas other serotypes,
namely nontyphoid Salmonella serotypes, can cause a
wide spectrum of diseases in humans and animals, such
as acute gastroenteritis, bacteremia, and extraintestinally
localized infections involving many organs Su et al.,
(2004).
Antimicrobial resistance has emerged in the past few
years as a major problem and many programs have been
set up for its surveillance in human and veterinary
medicine. These programs are aimed mainly at human
pathogens, agents of zoonoses, and indicator bacteria of
the normal intestinal flora from animals (LANZ). The
potential transfer of antibiotic resistance from animals to
humans through the use of antibiotics in animal
production has been implicated as a cause of treatment
failure, prolongation of illness and death, and increased
costs of treatment (Kelly) and (Kolar).
Hence, there is an urgent need for alternative
antibacterial strategies, and thus this condition has led to
a re-evaluation of the therapeutic use of new chemical
modifications and derivatives against most notorious
bacterial agents those responsible for serious drastic
infections.
Materials used
1- Tryptic Soy Broth for culturing and refreshment of
identified bacterial isolates in order to detect the effect of
modified antibacterial agents upon.
2-Muller Hinton agar that's used to test the effect of
modified antibacterial agents.
3- Identified bacterial isolates (Pseudomonasaeruginosa,
Staphylococcus aurous, Aeromonashydrophila,
Pasteurellamultocida, Ornithobacteriumrhinotracheale,
Escherichia coli, Salomnellatyphimurium and Salmonella
enterritidis).
4- Antibiotic reference like neomycin, doxycycline,
chloramphenicol, Cefixitin, streptomycin, Ampicillin,
penicillin, ofloxacin, erythromycin, amoxicillin and
tetracycline.
Methods:
1-detection of the bacterial count after 24hrs growth
according to Wiegand the isolated and identified bacterial
isolates under test was cultured in Tryptic Soy Broth (TSB)
for 24 hrs., then the concentration of bacterial cells in 1ml
medium was measured using spectrophotometer at 660
nm so as to adjust the concentration to 1x108colony
forming unit (CFU) per 1ml. So as to reach this bacterial
concentration sterile TSB used for dilution of the
concentrated bacterial isolates. from the adjusted 1
108CFU /l-11ml was taken and separated on the surface of
Muller Hinton agar plates and the excess decanted away
then the plates are left to dry at 40 0C in the incubator for
20 minutes. Wells made in agar plates by using the wide
end of a blunted sterile Pasteur pipette, inserting it and
twisting it slightly to remove the plug of agar.
Alternatively, cork borers sterilized with alcohol may be
used. A mounted needle or a pair of forceps, sterilized by
flaming in alcohol, may be required to remove the agar
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211
then, these wells were numbered with relevant numbers
of the utilized chemical substances furthermore, these
wells were filled with 120µl of the utilized modified
chemicals originated from antibiotics and then incubated
at 370C for 24-48hrs
The zone of inhibition traced on to an acetate grid with
felt pen and measured against squared or graph paper or
measured ruler to determine its size BSAC (2003).
Wiegand, I.; Hilpert, K.and Hancock, R. E. (2008): Agar and
broth dilution methods to determine the minimal
inhibitory concentration (MIC) of antimicrobial
substances.
Journal of Nature Protocols| 3(2):163-175. BSAC (2003)
Disc Diffusion Method for Antimicrobial Susceptibility
Testing Version 2.1.5 pages 1-41.
The results In Vitro Screening of samples for
antimicrobial activity after solubility in DMSO solvent on
cold show:
The efficacy of the novel derivatives of quinoxaline ring
from (.i) to (.xx) with concentration of 100% against
Pseudomonas aeruginosa as gram negative bacteria was
eminent as follows: No. (.i) (15mm), No. (.ii) (15mm), No.
(.iv) (13mm), No. (.v) (11mm), No. (.vi) (13mm), No. (.vii)
(10mm), No. (.viii) (12mm), No. (.ix) (12mm), No. (.x)
(15mm), No. (.xi) (15mm), No. (.xii) (18mm), No. (.xiii)
(8mm), No. (.xiv) (13mm), No. (.xv) (10mm), No. (.xvi)
(10mm), No. (.xviii) (12mm) and No. (.xx) (13mm) .As it is
shown in (figure 2).
Was noticeable when being compared with the
diminished efficacy of the original sulfaquinoxaline which
contains quinoxaline ring as well as resistance to most
antibiotics as Amoxicillin, Ampicillin, Penicillin,
Tetracycline, Neomycin, Chloramphenicol, Ofloxacin,
Doxycycline, and Erythromycin. As it is shown in (figure
3).
Furthermore, there located illustrious efficacy against
staphylococcus aureus as gram positive bacteria was
eminent as follows with derivatives No. (.i) (11mm), No.
(.ii) (17mm), No. (.iii) (10 mm), No. (.iv) (12mm), No. (.vi)
(10mm), No. (.vii) (15mm), No. (.viii) (10mm), No. (.ix)
(10mm), No. (.x) (18mm), No. (.xi) (7mm), No. (.xiii)
(8mm), No. (.xv) (7mm), No. (.xvi) (11mm), No. (.xviii)
(8mm) and No. (.xx) (8mm) .As it is shown in (figure 2).
This compared with the absence of efficacy of the original
sulfaquinoxaline which contains quinoxaline ring and
resistance to Amoxicillin, Tetracycline, Ofloxacin,
Doxycycline and Penicillin. As it is shown in (figure 3).
While, the efficacy of these derivatives was prominent
against Aeromonas hydrophila as gram negative bacteria
was eminent as follows: No. (.i) (13 mm), No. (.ii) (15mm),
No. (.iii) (20 mm), No. (.iv) (22mm), No. (.v) (23mm), No.
(.vi) (18mm), No. (.vii) (20mm), No. (.viii) (22mm), No.
(.ix) (15mm), No. (.x) (13mm), No. (.xi) (14mm), No. (.xii)
(17mm), No. (.xiii) (18mm), No. (.xiv) (12mm), No. (.xv)
(10mm), No. (.xvi) (15mm) and No. (.xiii) (10mm)
furthermore, it was apparent that derivatives (.iii), (.v),
(.vii) and (.viii) expressed synergistic effect. As it is shown
in (figure 2).
The effect of these products are promising with regard to
resistance to sulfaquinoxaline and most antibiotics as
Amoxicillin, ampicillin, Tetracycline, Ofloxacin,
Doxycycline, Penicillin, Chloramphenicol and
Erythromycin that have no effect against pseudomonas
aeruginosa. As it is shown in (figure 3).
Concerning efficacy against pasteurella multocida as
gram negative bacteria was eminent as follows on the
contrary to the wider range of efficacy of most derivatives
on previous bacterial species only No. (.vi) (7mm), No.
(.vii) (10mm) and No. (.viii) (10mm).
As it is shown in (figure 2).gave efficacy but still
conspicuous when compared with the resistance to
sulfaquinoxaline, Amoxicillin, Ampicillin, Penicillin,
Tetracycline, Neomycin, Streptomycin, Ofloxacin,
Doxycycline and Erythromycin. As it is shown in (figure
3).
Focusing upon the efficacy upon Ornithobacterium
rhinotracheale as gram negative bacteria was eminent as
follows only derivative's. No. (.i) (15mm), No. (.ii)
(18mm), No. (.iii) (10mm), No. (.iv) (12mm), No. (.vi)
(10mm), No. (.vii) (10mm), No. (.viii) (7mm), No. (.ix)
(10mm), No. (.x) (10mm), No. (.xii) (12mm), No. (.xiii)
(10mm), No. (.xv) (10mm) and No. (.xvi) (7mm). As it is
shown in (figure 2).
When compared with the no effect of quinoxaline ring
and most antibiotics as Amoxicillin, Ampicillin,
Chloramphenicol, Tetracycline, Neomycin, Streptomycin,
Ofloxacin, Doxycycline and Erythromycin. As it is shown
in (figure 3).
Interpreting the effect against Escherichia coli as gram
negative bacteria was eminent as follows only
derivative's .No. (.i) (20mm), No. (.iii) (14mm), No. (.v)
(25mm), No. (.vii) (15mm), No. (.viii) (25mm), No. (.ix)
(15mm), No. (.xv) (10mm), No. (.xvi) (10mm) and No.
(.xviii) (10mm).
As it is shown in (figure 2). with regard to
sulfaquinoxaline which expressed less effect against
E.coli as well as Ampicillin, Penicillin, Tetracycline,
Neomycin, Streptomycin, Ofloxacin, Doxycycline,
Erythromycin and Chloramphenicol. As it is shown in
(figure 3).
Concerning the susceptibility of Salmonella typhimurium
as gram negative bacteria was eminent as follows to
most derivatives' only products .No. (.i) (10mm), No. (.ii)
(16mm), No. (.iv) (15mm), No. (.vi) (12mm), No. (.viii)
(15mm), No. (.ix) (10mm) and No. (.xii) (17mm). As it is
shown in (figure 2). when compared with
sulfaquinoxaline which gave no efficacy against
Salomnella typhimurium as well as Ampicillin,
Amoxicillin, Penicillin, Neomycin, quinolones as
Int. J. Pharm. Sci. Rev. Res., 34(1), September – October 2015; Article No. 33, Pages: 205-213 ISSN 0976 – 044X
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Ofloxacin, Doxycycline, Erythromycin and
Chloramphenicol. As it is shown in (figure 3).
On the contrary to most tested bacterial species
Salmonella enteritidis as gram negative bacteria was
eminent as follows the lowest susceptible one only
derivative. No. (.ii) (10 mm)
As it is shown in (figure 2). Gave efficacy against it but it
still encouraging when compared with resistance to
sulfaquinoxaline, Ampicillin, Amoxicillin, Penicillin,
Neomycin, quinolones as Ofloxacin, Doxycycline,
Erythromycin, Chloramphenicol, Tetracycline and
Streptomycin. As it is shown in (figure 3).
CONCLUSION
The synthesized derivatives from the quinoxaline ring are
promising due to the salient effect against the tested
antibacterial species.
This mainly opens the door to evade the concurrent
problem of bacterial resistance but further investigations
are required to test and detect the pharmacokinetics
(absorption, distribution, metabolism, and excretion),
Toxic kinetics in animal's effective dosage, over dosage of
these valuable products.
Acknowledgement: We thank Prof. Dr. Gerhard Maas,
Institut für Organische Chemie, the University of Ulm,
Germany for assistance with NMR measurements.
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Source of Support: Nil, Conflict of Interest: None.
... An important strategy in fighting antibiotic resistance is the discovery, development of novel antibiotics, and increasing the efficacy of the antibiotic that is already in a clinical study [7]. In this context, α-aminophosphonates gained great interest by medicinal chemists because of their diverse biological and industrial applications such as antibacterial [8][9][10][11][12], anticancer [13][14][15], enzyme inhibitors [16,17], and chelating material [18][19][20][21] which made them a promising drug candidate for further optimization. These compounds are phosphorus analogs of naturally occurring α-amino acids and therefore, are considered promising in the field of drug discovery and development [22]. ...
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