Synthesis of 1-amidoalkyl-2-naphthols and oxazine derivatives with study of their antibacterial and antiviral activities

Article abstract of DOI:10.1007/s00044-012-0205-9

An efficient and direct protocol for the preparation of 1-amidoalkyl-2-naphthols 1a-f employing a multi-component, one-pot condensation reaction of 2-naphthol, heterocyclic aldehydes, and amides in the presence of anhydrous zinc chloride under solvent-free conditions is described. The thermal solvent-free offer advantages such as shorter reaction times, simple work-up and excellent yield. Ring closure of 1a-f gave the pyrazolyl- and indolyl oxazine derivatives 2a-f. On the other hand, the reaction of 2-naphthol, aldehydes, and ammonia solution gave the dipyrazolyl- and di-indolyl oxazine derivatives 3a,b. Some of the newly synthesized compounds showed promising antibacterial and anti-H₅N₁ activities.

Full text of DOI:10.1007/s00044-012-0205-9

MEDICINAL
CHEMISTRY
RESEARCH
Med Chem Res (2013) 22:2005–2013
DOI 10.1007/s00044-012-0205-9
ORIGINAL RESEARCH
Synthesis of 1-amidoalkyl-2-naphthols and oxazine derivatives
with study of their antibacterial and antiviral activities
•
Wael S. I. Abou-Elmagd Ahmed I. Hashem
Received: 30 April 2012 / Accepted: 10 August 2012 / Published online: 25 August 2012
Ó Springer Science+Business Media, LLC 2012
Abstract An efficient and direct protocol for the prepara-
tion of 1-amidoalkyl-2-naphthols 1a–f employing a multi-
component, one-pot condensation reaction of 2-naphthol,
heterocyclic aldehydes, and amides in the presence of
anhydrous zinc chloride under solvent-free conditions is
described. The thermal solvent-free offer advantages such as
shorter reaction times, simple work-up and excellent yield.
Ring closure of 1a–f gave the pyrazolyl- and indolyl oxazine
derivatives 2a–f. On the other hand, the reaction of
2-naphthol, aldehydes, and ammonia solution gave the
dipyrazolyl- and di-indolyl oxazine derivatives 3a,b. Some
of the newly synthesized compounds showed promising
antibacterial and anti-H5N1 activities.
processes, but also help to prevent the releasing of toxic
reaction residues into the environment.
One-pot multi-components condensation of 2-naphthol,
aromatic aldehydes, and amide derivatives has been used as a
practical synthetic routes toward 1-amidoalkyl-2-naphthols.
Several Lewis and Brønsted acids have been applied for this
transformation (Shaterian and Yarahmadi, 2008; Das et al.,
2007; Nagawade and Shinde, 2007; Shaterian et al., 2008;
Nagarapu et al., 2007; Mahdavinia et al., 2008; Hajipour
et al., 2009; Nandi et al., 2009; Mahdavinia et al., 2009;
Kantevari et al., 2007; Patil et al., 2007; Lei et al., 2009; Su
et al., 2008, Srihari et al., 2007; Kumar et al., 2009; and
Selvam et al., 2006). However, many of the reported meth-
ods suffer from one or more of the following drawbacks:
(i) low product yield, (ii) prolonged reaction time, (iii) the
use of large amount of reagents, (iv) the use of toxic reagent,
and (v) incompatibility with the green chemistry protocol.
Therefore, search for finding a protocol for the synthesis of
1-amidoalkyl-2-naphthols that are not associated with the
above disadvantages is still relevant.
Keywords 1-Amidoalkyl naphthol ꢀ Oxazine ꢀ
Zanamivir ꢀ Antibacterial ꢀ Anti-H5N1
Introduction
The development of simple synthetic routes to widely used
organic compounds using readily available reagents is one
of the main objectives of organic synthesis. Nitrogen het-
erocyclic compounds are of special interest. They consti-
tute an important class of natural and non-natural products,
many of which exhibit useful biological activities (Turgut
et al., 2007, Xu and Guo, 2004, Hashem et al., 2007).
Recently, the use of catalysts and reagents supported on
solid supports under solvent-free conditions has been
developed. Such reagents not only simplify the purification
1-Amidoalkyl-2-naphthol derivatives are of importance
because they can be converted by carbamate hydrolysis
into the biologically active 1- aminoalkyl-2-naphthol. The
latter products were shown to have hypotensive and brad-
¨
¨ ¨
ycardiac effects (Szatmari and Fulop, 2004).
The synthesis of 1,3-oxazines has attracted attention in
the past because of their potential as antibiotics, antitumor
agents, analgesics, and anticonvulsants. 1,3-Oxazines have
generated great interest as antipsychotic agents and as
possible effectors for serotonin and dopamine receptors. In
addition, benzo-1,3-oxazines are known to be biologically
active as anti-malarial, anti-anginal, anti-hypertensive, and
potent anti-rheumatic agents (Shen et al., 1999).
W. S. I. Abou-Elmagd (&) ꢀ A. I. Hashem
Chemistry Department, Faculty of Science,
Ain Shams University, Abassia, Cairo 11566, Egypt
e-mail: waelmagd97@yahoo.com
Zanamivir is a neuraminidase inhibitor used in the
treatment and prophylaxis of influenza caused by influenza
123

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Med Chem Res (2013) 22:2005–2013
A virus and influenza B virus (Soundararajan et al., 2009).
The presence of oxine ring with the presence of free
hydroxy, amino, amide, and carboxylic acid groups initi-
ated our interest to synthesize the amidoalkyl-2-naphthol
and oxazine ring containing one or more of these groups.
We wish to report ‘‘Herein’’ a new, efficient and simple
method for the synthesis of 1-amidoalkyl-2-naphthols, pyr-
azolyl- and indolyl-1,3-oxazine derivatives from 2-naphthol,
heterocyclic aldehydes and amides in the presence of anhy-
drous zinc chloride as catalyst under solvent-free conditions.
Selected examples of the compounds obtained were evalu-
ated for their antibacterial and antiviral activities.
Results and discussion
Chemistry
I,3-Diphenylpyrazol-4-carboxaldehyde and/or indole-3-
carboxaldehyde were condensed with 2-naphthol and
amide (urea, acetamide, benzamide) in the presence of
anhydrous zinc chloride at 80–100 °C for 1 h under sol-
vent-free conditions (Scheme 1). The reaction proceeded
smoothly and gave the corresponding 1-amidoalkyl-2-
naphthols 1a–f as sole products in 90 % yield.
On heating 1a–f in absolute ethanol, ring closure
occurred with the formation of the corresponding 1,3-
oxazine derivatives 2a–f. It is to be mentioned that, this
ring clousure was also affected by refluxing 1a–f with P2O5
in dry toluene (cf., ‘‘Experimental’’ section).
The structures of the 1-amidoalkyl-2-naphthols and 1,3-
oxazine derivatives were inferred from their analytical and
spectral data (cf., ‘‘Experimental’’ section).
The infrared spectra of 1a–f revealed the disappearance
of the characteristic C=O bands of the aldehydes and
absorption bands corresponding to NH groups were
observed between 3,219 and 3,321 cm-1, as well as, amide
carbonyl group at 1,640–1,650 cm-1. The NCH proton
Scheme 1 Synthesis of
1-amidoalkyl-2-naphthols and
1,3-oxazine derivatives
OH
O
O
H
anhyd.
ZnCl2
+
R
+
X
NH2
OH
X
NH
Ph
R= NH2 , CH3 , Ph
,
X=
R
O
N
1
N
N
H
Ph
reflux
Absolute
Ethanol
O
X
N
R
2
a, X=1,3-diphenyl pyrazolyl
b, X= 1,3-diphenyl pyrazolyl
c, X=1,3-diphenyl pyrazolyl
d, X= indolyl
e, X= indolyl
f, X= indolyl
R= NH2
R= CH3
R= Ph
R= NH2
R= CH3
R= Ph
123

Med Chem Res (2013) 22:2005–2013
2007
singlet between 8.02 and 8.41 ppm were observed in the
1H-NMR spectra in case of 1a–f and the disappearance of
NH and OH groups in case of 2a–f (Scheme 1).
data which show the disappearance of the characteristic
C=O bands of the aldehydes and OH band of 2-naphthol
and the appearance of the saturated oxazine NH at
3,127–3,234 cm-1 (cf., ‘‘Experimental’’ section)
We believe that the formation of 1a–f is similar to the
Hoesch reaction in which a chloroimine attacks activated
phenols. So, it is possible that the aldehyde reacts firstly
with the amide to give the imine X–CH=N–CO–R. The
latter intermediate has an a,b—unsaturated carbonyl moi-
ety, and hence can attack position -1 in 2-naphthol via its
highly deficient carbon (cf. Scheme 2)
Biological evaluation
Cytotoxicity and antiviral activity
On the other hand, when 1,3-diphenyl-4-carboxaldehyde
and/or indole-3-carboxaldehyde were allowed to react with
2-naphthol in ammonia solution according to Mannich type
aminoalkylation by applying classical Betti’s reaction
(Scheme 3; Betti, 1942). The reaction proceeded smoothly
and gave the corresponding dipyrazolyl oxazine and di-
indolyl oxazine derivatives 3a,b. The structures of the new
compounds obtained have been clarified by spectroscopic
Selective compounds were evaluated for their cytotoxicity
using African green monkey kidney (Vero) cells using the
3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bro-
mide (MTT) method and were evaluated for their antiviral
activity using cytopathogenicity (CPE) assay against avian
influenza virus (H5N1) taking Zanamivir as a control. The
results of the assay are summarized in Table 1 and illustrated
by Fig. 1. All compounds except 1f showed better
Scheme 2 A proposed pathway
for the formation of
1-amidoalkyl-2-naphthols and
1,3-oxazine derivatives
O
O
O
H
2-naphthol
+
R
X
C
H
N
X
ZnCl2
NH2
R
OH
CH
X
N
-:O:
R
..
+H+
OH
OH
OH
X
NH
X
N
R
O
R
-H2O
O
X
N
R
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Med Chem Res (2013) 22:2005–2013
H
Scheme 3 A proposed pathway
for the formation of diheteryl
1,3-oxazine derivatives
X
N
X
OH
O
O
H
NH3 / MeOH
+
2
X
3
..-
X
NH
..
CH
OH
O
H
2-naphthol
NH3
+
X
X
C
H
NH
+H+
X
NH2
X
N
C
H
X
O
H
CH
CH
X
..
OH
OH
..
-H+
+H+
Ph
H
N
X
X
; b,
X= a,
N
O
N
N
H
Ph
3
cytotoxicity than the control Zanamivir. Among them,
compounds 1c, 2a, and 3a showed cytotoxicity against cell
line below a concentration of 0.1 lM and 1a, 1b, 1e, 2d, 2f,
and 3b showed cytotoxicity below a concentration of
0.2 lM. The antiviral results indicated that compounds 1c
and 1f were the most active compounds with EC50 values
0.018 and 0.019, respectively.
generally more potent in antiviral activity. Finally, the
higher effective compound than the control Zanamivir was
1f.
Most of the newly synthesized compounds were
screened for their antimicrobial activity against variety of
human pathogenic bacteria; the results of the assay are
summarized in Table 2.
The preliminary structure–activity relationships sug-
gested that compounds with free amino, amide, or hydroxyl
groups were generally more potent in cytotoxicity than the
cyclic oxazine ring lacking one or more of these groups.
But the oxazine ring having a pyrazole moiety was
The results of the assay showed that the amidoalkyl
naphthols 1a, 1b, 1c, and 1e gave high activity (Inhibition
zone) and the other compounds showed moderate activi-
ties. All compounds showed no activity towards P. aero-
ginosa, E.coli, and Proteus Microorganisms.
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Med Chem Res (2013) 22:2005–2013
2009
Conclusion
Table 1 Cytotoxicity and antiviral activity
Compound
LD50
(lM/ml)
EC50 (lM/ml)
Avian Influenza
Virus (H5N1)
Therapeutic
index
A new, efficient, and simple method for the synthesis of
1-amidoalkyl-2-naphthols, pyrazolyl and indolyl-1,3-oxa-
zine derivatives from 2-naphthol, heterocyclic aldehydes,
and amides in the presence of anhydrous zinc chloride as
catalyst under solvent-free conditions is reported. On the
other hand, the reaction of 2-naphthol, aldehydes, and
ammonia solution gave the dipyrazolyl- and di-indolyl
oxazine derivatives. Some of the newly synthesized com-
pounds showed promising antibacterial and anti-H5N1
activities. Compounds with free amino, amide, or hydroxyl
groups were generally more potent in cytotoxicity and
antibacterial activities than the cyclic oxazine ring lacking
these groups. The oxazine ring having pyrazole moiety was
generally more potent in antiviral activity.
1a
0.171
0.151
0.098
0.217
0.197
0.307
0.083
0.125
0.104
0.089
0.122
0.232
0.025
0.021
0.018
0.031
0.022
0.019
0.0148
0.061
0.049
0.045
0.168
0.015
6.84
1b
7.19047619
5.444444444
7
1c
1d
1e
8.954545455
16.15789474
5.608108108
2.049180328
2.12244898
1.977777778
0.726190476
15.46666667
1f
2a
2d
2f
3a
3b
Zanamivir
Experimental
Melting points are measured on an electrothermal melting
point apparatus. Elemental analyses were carried out at the
Microanalytical Unit, Cairo University. The IR spectra
were measured on a Unicam SP-1200 spectrometer using
1
KBr Wafer technique. The H-NMR spectra were mea-
sured in DMSO-d6 on a Varian plus instrument (300 MHz).
Mass spectra were recorded on a Shimadzu GC–MS
QP- 1000EX instrument operating at 70 eV.
Chemistry
General procedure for the preparation of 1-amidoalkyl-
2- naphthols 1a–f
A mixture of aldehyde (1 mmol), 2-naphthol (1 mmol),
amide (1.3 mmol), and anhydrous ZnCl2 (0.25 mmol) were
heated at 80–100 °C for 1 h. After completion of the
reaction (monitored by TLC), the reaction mixture was
diluted with water, and the resulting solid product was
collected by filtration and recrystallized from 30 % aque-
ous ethanol to give 1-amidoalkyl-2-naphthols 1a–f.
Fig. 1 Cytotoxicity and antiviral activity
Table 2 Antibacterial activity (as inhibition zone in mm diameter)
Sample no./
microorganism
1a 1b 1c 1d 1e 1f 2a 2d 2f 3a 3b
K. pneumonia 11 18 14 11
–
–
–
–
–
–
–
–
11 13 11 11 12
1-((1,3-Diphenyl-1H-pyrazol-4-yl)(2-hydroxynaphthalen-
1-yl)methyl)urea 1a
P. aeroginosa
E. coli
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
Proteus
0
–
Colorless crystals; m.p: 168–170 °C, yield 85 %. IR (KBr)
(mmax, cm-1): 3465–3348(br.) (OH), 3321, 3237, 3219
S. Aureus
B. subtilis
24
–
–
20 11
11 11
20
17 11 12 12 14 11 13
1
(NH, NH2), 1640 (C=O). H-NMR (DMSO-d6): dH (ppm)
Standard reading using gentamicin antibiotic 0.042 lM/ml. The
inhibition zone exhibited by gentamicin 0.042 lM/ml in mm diam-
eter. K. pneumonia 19 mm; P. aeurginosa 27 mm; E. coli 29 mm;
P. mirabilis 25 mm; S. aureus 28 mm; and B. subtilus 21 mm
6.95–7.80 (m, 17H, ArH), 7.84 (br.s, 1H, OH, exchange-
able with D2O), 8.43 (s, 1H, CH), 8.71 (br.s, 2H, NH2,
exchangeable with D2O), 9.10 (br.s, 1H, NH, exchangeable
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Med Chem Res (2013) 22:2005–2013
with D2O). MS, m/z (%): 434 (M.?, 17), 285 (31), 246 (52),
190 (63), 161 (57), 110 (37), 77 (100). Anal. Calcd. For
C27H22N4O2 (434): C, 74.64; H, 5.10; N, 12.89. Found: C,
75.12; H, 5.33; N, 12.76.
1H, NH, exchangeable with D2O). MS, m/z (%): 330 (M.?,
7), 287(69), 143(37), 98(57), 81 (100), 80 (32), 63 (23).
Anal. Calcd. For C21H18N2O2 (330): C, 76.34; H, 5.49; N,
8.48. Found: C, 76.69; H, 5.24; N, 8.80.
N-((1,3-Diphenyl-1H-pyrazol-4-yl)(2-hydroxynaphthalen-
1-yl)methyl)acetamide 1b
N-((2-Hydroxynaphthalen-1-yl)(1H-indol-3-
yl))methyl)benzamide 1f
Colorless crystals; m.p: 230–231 °C, yield 90 %. IR (KBr)
(mmax, cm-1): 3470–3380(br.) (OH), 3250 (NH), 1649
(C=O). 1H-NMR (DMSO-d6): dH (ppm) 2.31 (s, 3H, CH3),
6.90–7.77 (m, 17H, ArH), 7.81 (br.s, 1H, OH, exchange-
able with D2O), 8.43 (s, 1H, CH), 9.14 (br.s, 1H, NH,
exchangeable with D2O). MS, m/z (%): 433 (M.?, 25),
391(50),285 (27), 246 (34), 190 (50), 161 (43), 110 (39), 77
(100). Anal. Calcd. For C28H23N3O2 (433): C, 77.58; H,
5.35; N, 9.69. Found: C, 77.72; H, 5.20; N, 9.86.
Colorless crystals; m.p: 230–232 °C, yield 91 %. IR (KBr)
(mmax, cm-1): 3477–3393(br.) (OH), 3312, 3250 (NH),
1650 (C=O). H-NMR (DMSO-d6): dH (ppm) 7.04–7.82
(m, 17H, ArH ? NHindole), 7.83 (br.s, 1H, OH,
exchangeable with D2O), 8.08 (s, 1H, CH), 9.02 (br.s, 1H,
NH, exchangeable with D2O). MS, m/z (%): 392 (M.?, 43),
287(56), 285 (34), 240 (32), 192 (35), 144 (65), 110 (30),
81 (56), 77 (100). Anal. Calcd. For C26H20N2O2 (392): C,
79.57; H, 5.14; N, 7.14. Found: C, 79.29; H, 5.21; N, 7.06.
1
N-((1,3-Diphenyl-1H-pyrazol-4-yl)(2-hydroxynaphthalen-
1-yl)methyl)benzamide 1c
General procedure for the cyclization of compounds
1a–f; formation of 1,3-oxazine derivatives 2a–f
Colorless crystals; m.p: 275–278 °C, yield 89 %. IR (KBr)
(mmax, cm-1): 3420–3350(br.) (OH), 3226 (NH), 1652
(C=O). 1H-NMR (DMSO-d6): dH (ppm) 7.04–7.82 (m,
22H, ArH), 7.88 (br.s, 1H, OH, exchangeable with D2O),
8.41 (s, 1H, CH), 9.15 (br.s, 1H, NH, exchangeable with
D2O). MS, m/z (%): 495(M.?, 12), 391(67), 285 (34), 246
(46), 190 (54), 161 (64), 110 (30), 77 (100). Anal. Calcd.
For C33H25N3O2 (495): C, 79.98; H, 5.05; N, 8.48. Found:
C, 79.59; H, 5.23; N, 8.66.
A solution of 1a–f in absolute ethanol was heated for 1 h.
and then left to cool. The solid obtained was filtered off and
recrystallized from absolute ethanol to give the oxazine
derivatives 2a–f.
The same products 2a–f were obtained when a solution
of 1a–f (1 mmol) and P2O5 (1.1 mmol) in dry toluene were
refluxed for 1 h. The whole mixture was concentrated and
left to cool. The precipitated solid was filtered off, washed
with water, dried, and recrystallized from ethanol.
1-((2-Hydroxynaphthalen-1-yl)(1H-indol-3-
yl))methyl)urea 1d
1-(1,3-Diphenyl-1H-pyrazol-4-yl)-1H- naphtho[1,2-
e][1,3]oxazin-3-amine 2a
Colorless crystals; m.p: 142-144 °C, yield 88 %. IR (KBr)
(mmax, cm-1): 3495–3358(br.) (OH), 3300, 3280, 3267
(NH, NH2), 1647 (C=O). H-NMR (DMSO-d6): dH (ppm)
Pale yellow crystals; m.p: 125–128 °C, yield 59 %. IR
1
1
(KBr) (mmax, cm-1): 3227, 3210 (NH2), 1619 (C=N). H-
7.05–7.67 (m, 12H, ArH ? NHindole), 7.75 (br.s, 1H, OH,
exchangeable with D2O), 8.16 (s, 1H, CH), 8.95 (br.s, 2H,
NH2, exchangeable with D2O), 9.08 (br.s, 1H, NH,
exchangeable with D2O). MS, m/z (%): 331 (M.?, 34), 288
(48), 271 (65), 190 (32), 160 (50), 144 (100), 75 (50), 57
(88). Anal. Calcd. For C20H17N3O2 (331): C, 72.49; H,
5.17; N, 12.68. Found: C, 72.78; H, 5.08; N, 12.97
NMR (DMSO-d6): dH (ppm) 4.52 (br.s, 2H, NH2,
exchangeable with D2O), 5.73 (s, 1H, CH), 6.90–7.67 (m,
17H, ArH). MS, m/z (%): 416 (M.?, 5), 286(43), 245 (54),
191 (47), 160 (73), 110 (30), 77 (100). Anal. Calcd. For
C27H20N4O (416): C, 77.87; H, 4.84; N, 13.45. Found: C,
77.52; H, 5.13; N, 13.76.
1-(1,3-Diphenyl-1H-pyrazol-4-yl)-3-methyl-1H-
naphtho[1,2-e][1,3]oxazin 2b
N-((2-Hydroxynaphthalen-1-yl)(1H-indol-3-
yl))methyl)acetamide 1e
Pale yellow crystals; m.p: 138–139 °C, yield 51 %. IR
(KBr) (mmax, cm-1): 3059, 3037 (CHaromatic), 2889, 2836,
2758 (CHaliphatic), 1622 (C=N). H-NMR (DMSO-d6): dH
(ppm) 1.93 (s, 3H, CH3), 5.43 (s, 1H, CH), 7.19–7.82 (m,
17H, ArH). MS, m/z (%): 415 (M.?, 9), 390(43), 286 (57),
246 (30), 191 (56), 161 (23), 112 (19), 77 (100), 57 (24).
Colorless crystals; m.p: 180–182 °C, yield 92 %. IR (KBr)
(mmax, cm-1): 3495–3350(br.) (OH), 3243, 3220 (NH),
1647 (C=O). H-NMR (DMSO-d6): dH (ppm) 2.19 (s, 3H,
CH3), 7.08–7.93 (m, 12H, ArH ? NHindole), 7.58 (br.s, 1H,
1
1
OH, exchangeable with D2O), 8.02 (s, 1H, CH), 8.97 (br.s,
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Med Chem Res (2013) 22:2005–2013
2011
Anal. Calcd. For C28H23N3O (415): C, 80.94; H, 5.09; N,
10.11. Found: C, 80.70; H, 5.36; N, 9.89.
separated out. The crude crystals were filtered off, washed
with cold MeOH, and recrystallized from absolute ethanol
to give the diheteryl-1,3-oxazine derivatives 3a,b.
1-(1,3-Diphenyl-1H-pyrazol-4-yl)-3-phenyl-1H-
naphtho[1,2-e][1,3]oxazin 2c
1,3-Bis(1,3-diphenyl-1H-pyrazol-4-yl)-2,3-dihydro-1H-
naphtho[1,2-e][1,3] oxazine 3a
Pale yellow crystals; m.p: 142–143 °C, yield 59 %. IR
(KBr) (mmax, cm-1): 3038, 3021 (CHaromatic), 2878 (CHali-
phatic), 1620 (C=N). H-NMR (DMSO-d6): dH (ppm) 5.77
(s, 1H, CH), 7.19–7.82 (m, 22H, ArH). MS, m/z (%):
477(M.?, 21), 258(32), 219 (63), 143 (78), 77 (100), 57
(30). Anal. Calcd. For C33H23N3O (477): C, 83.00; H, 4.85;
N, 8.80. Found: C, 82.79; H, 5.03; N, 8.66.
Creamy powder; m.p: 130–132 °C, yield 88 %. IR (KBr)
(mmax, cm-1):3127(NH), 3054, 3034, 3019(CHaromatic),2863,
2836, 2784 (CHaliphatic). 1H-NMR (DMSO-d6): dH (ppm) 4.32
(br.s, 1H, NH, exchangeable with D2O), 5.71 (s, 1H, CH–N),
6.48 (s, 1H, N–CH-O), 6.89–7.82 (m, 28H, ArH). MS, m/
z (%): 621 (M.?, 26), 519 (26), 451 (42), 305 (31), 285 (32),
247(47), 235(26), 190(63), 162(58), 110(36), 84(47), 77(100),
69(84). Anal. Calcd. For C42H31N5O (621): C, 81.14; H, 5.03;
N, 11.26. Found: C, 81.49; H, 4.88; N, 11.50.
1
1-(1H-indol-3-yl)-1H-naphtho[1,2-e][1,3]oxazin-
3-amine 2d
Pale yellow crystals; m.p: 107–110 °C, yield 45 %. IR (KBr)
(mmax, cm-1): 3302, 3227, 3210 (NH, NH2), 1620 (C=N). 1H-
NMR (DMSO-d6): dH (ppm) 4.39 (br.s, 2H, NH2, exchange-
able with D2O), 4.91 (s, 1H, CH), 6.84–7.71 (m, 12H,
Ar ? NHindole). MS, m/z (%): 313 (M.?, 3), 98 (57), 97 (22),
80 (30), 81 (100), 63 (23). Anal. Calcd. For C20H15N3O (313):
C, 76.66;H, 4.82;N,13.41.Found:C,76.97;H,4.98;N,13.76.
1,3-Di(1H-indol-3-yl)-2,3-dihydro-1H-naphtho[1,2-e][1,3]
oxazine 3b
Creamy powder; m.p: 190–192 °C, yield 80 %. IR (KBr)
(mmax, cm-1): 3234, 3169 (NH), 3044 (CHaromatic), 2980,
1
2935, 2989 (CHaliphatic). H-NMR (DMSO-d6): dH (ppm)
4.02 (br.s, 1H, NH, exchangeable with D2O), 5.29 (s, 1H,
CH–N), 6.21 (s, 1H, N–CH–O), 7.11–7.92 (m, 18H,
ArH ? 2NHindole). MS, m/z (%): 415 (M.?, 9), 360 (33),
216 (33), 200 (50), 186 (33), 172(44), 159(38), 144(100),
115(55), 98(50), 89(83), 73(61), 57(89). Anal. Calcd. For
C28H21N3O (415): C, 80.94; H, 5.09; N, 10.11. Found: C,
81.09; H, 5.28; N, 10.34.
1-(1H-indol-3-yl) -3-methyl-1H-naphtho[1,2-e]
[1,3]oxazin 2e
Pale yellow crystals; m.p: 122–124 °C, yield 52 %. IR
1
(KBr) (mmax, cm-1): 3243 (NH), 1618 (C=N). H-NMR
(DMSO-d6): dH (ppm) 1.96 (s, 3H, CH3), 4.52 (s, 1H, CH),
7.08–7.64 (m, 12H, Ar ? NHindole). MS, m/z (%):
312(M.?, 15), 290(42), 195 (51), 143 (100), 116 (67), 57
(54). Anal. Calcd. For C21H16N2O (312): C, 80.75; H, 5.16;
N, 8.97. Found: C, 80.49; H, 5.27; N, 8.83.
Biology
Cytotoxicity assay
1-(1H-indol-3-yl) -3-phenyl-1H-naphtho
[1,2-e][1,3]oxazin 2f
The stock samples were diluted with Dulbecco’s Modified
Eagle’s Medium (DMEM) to desired concentrations. Stock
solutions of the test compounds were prepared in DMSO at
a concentration of 10 % in D2O. The cytotoxic activity of
the synthetic compounds were tested in African green
monkey kidney (Vero) cells using the 3-(4,5-dimethylthi-
azol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) method
(Mossman, 1983) with minor modification. In brief, the
cells were seeded in 96 well-plates (100 ll/well at a den-
sity of 3 9 105 cells/ml) and treated with various con-
centrations of the sample solutions. After 24 h, cells were
washed with sterile phosphate buffer (PBS) three times and
the supernatant was discarded. MTT solution (20 ll of
5 mg/ml) was added to each well and incubated at 37 °C
for 4 h then the medium was aspirated. In each well, the
formed Formosan crystals were dissolved with 200 ll of
acidified 2-propanol (0.04 M HCl in absolute 2-propanol).
Pale yellow crystals; m.p: 131–134 °C, yield 51 %. IR (KBr)
(mmax, cm-1): 3307 (NH), 1623 (C=N). 1H-NMR (DMSO-d6):
dH (ppm) 5.81 (s, 1H, CH), 7.18–7.92 (m, 17H, Ar ? NHin-
dole). MS, m/z (%): 374(M.?, 13), 296(25), 219 (59), 143 (82),
76 (100), 57 (76). Anal. Calcd. For C26H18N2O (374): C,
83.40; H, 4.85; N, 7.48. Found: C, 83.19; H, 5.03; N, 7.19.
General procedure for the preparation of diheteryl-1,3-
oxazine derivatives 3a,b
Aldehyde (2 mmol) and (15 ml) ammonia solution (33 %)
were added to a solution of 2-naphthol (1 mmol) in abso-
lute MeOH (30 ml). The mixture was left to stand at 0 °C
for 2 days, whereby a crystalline product was formed and
123

2012
Med Chem Res (2013) 22:2005–2013
An absorbance of Formosan was detected by a dual
wavelength UV spectrometer at 540 nm with 620 nm ref-
erence wavelength. The percentage of cytotoxicity com-
pared to the untreated cells was determined with the
following equation:
aureus] were used. The cultures of bacteria were main-
tained in their appropriate agar slants at 4 °C throughout
the study and used as stock cultures. The inhibition zone
was measured for each of the tested compounds taking
Gentamycin as a reference.
ðAbsorbance of cell without treatment ꢁ Absorbance of cell with treatmentÞ ꢂ 100
% Cytotoxicity ¼
Absorbance of cell without treatment
Acknowledgments The authors are thankful to Dr. Alaa R.I. Morsy,
Central Laboratory for Evaluation of Veterinary Biologics, Agricul-
tural Research Center, Abassia, Cairo, Egypt and Dr. Madeha O.I.
Ghobashy, Microbiology Department, Faculty of Science, Ain Shams
University for biological investigations.
The plot of % cytotoxicity versus sample concentration
was used to calculate the concentration which exhibited
50 % cytotoxicity (LD50).
Antiviral assay
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