Substituted 3-((Z)-2-(4-nitrophenyl)-2-(1H-tetrazol-5-yl) vinyl)-4H-chromen-4-ones as novel anti-MRSA agents: Synthesis, SAR, and in-vitro assessment

Article abstract of DOI:10.1016/j.bmcl.2008.07.007

In search for a new antibacterial agent with improved antimicrobial spectrum and potency, we designed and synthesized a series of novel 3-((Z)-2-(4-nitrophenyl)-2-(1H-tetrazol-5-yl) vinyl)-4H-chromen-4-ones 7a-h by convergent synthesis approach. All the synthesized compounds were assayed for their in-vitro antibacterial activities against gram-negative and gram-positive bacteria.

Full text of DOI:10.1016/j.bmcl.2008.07.007

Bioorganic & Medicinal Chemistry Letters 18 (2008) 4678–4681
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Substituted 3-((Z)-2-(4-nitrophenyl)-2-(1H-tetrazol-5-yl)
vinyl)-4H-chromen-4-ones as novel anti-MRSA agents: Synthesis, SAR,
and in-vitro assessment
Santosh D. Diwakar a,c, Sachin S. Bhagwat b, Murlidhar S. Shingare c, Charansing H. Gill c,
*
a Drug Discovery Research, Chemistry, Wockhardt Research Centre, D-4, MIDC, Chikalthana, Aurangabad 431 210, India
b Drug Discovery Research, Biology, Wockhardt Research Centre, D-4, MIDC, Chikalthana, Aurangabad 431 210, India
c Department chemistry, Dr. Babasaheb Ambedkar Marathwada University, Aurangabad-431004, India
a r t i c l e i n f o
a b s t r a c t
Article history:
In search for a new antibacterial agent with improved antimicrobial spectrum and potency, we designed
and synthesized a series of novel 3-((Z)-2-(4-nitrophenyl)-2-(1H-tetrazol-5-yl) vinyl)-4H-chromen-4-
ones 7a–h by convergent synthesis approach. All the synthesized compounds were assayed for their
in-vitro antibacterial activities against gram-negative and gram-positive bacteria. The preliminary struc-
ture-activity relationship, to elucidate the essential structure requirements for the antimicrobial activity
that results into anti-MRSA (methicillin-resistant S. aureus) potential, has been described. Amongst the
synthesized compounds 7d, 7e, 7f and 7h were found to possess activity against methicillin-resistant
S. aureus in addition to the activity against other bacterial strains such as E. faecalis, S. pneumoniae, and
E. coli.
Received 1 April 2008
Revised 13 June 2008
Accepted 3 July 2008
Available online 9 July 2008
Keywords:
Chromones
Tetrazole
Anti-MRSA
Convergent
E. faecalis
Ó 2008 Elsevier Ltd. All rights reserved.
Owing to the emergence of multi-drug-resistant strains in re-
cent decades, microbial diseases have become more complex to
tackle as compared to the first half of the last century. To combat
such microbial infections, various synthetic and semi-synthetic
antimicrobial drugs have been discovered and used in the clinical
practice. In spite of significant developments in the antimicrobial
therapy, problems such as resistance development, spectrum of
activity, potency and toxicity remain unresolved for most of the
available antimicrobial drugs. Hence, the discovery and develop-
ment of novel antimicrobial agents with optimized pharmacologi-
cal profile together with increased activity towards resistant
strains are highly desirable.
Tetrazole is a class of heterocycles with wide range of applica-
tions that is receiving considerable attention.6 This functionality
has been frequently used as metabolically stable surrogates for car-
boxylic acid group as well as lipophilic spacers.7 Attempts to re-
place the carboxylic acid group with a tetrazole ring have been
made with chromones.8a,b Since 1H-tetrazole generally shows
acidity comparable with Carboxylic acid (Fig. 1), 3-(1H-tetrazole-
5-yl)-chromone derivatives 2 (R = H; pKa = 5.85), which proved to
be stronger acid than 1 (R = H; pKa = 8.85), were found to be active,
not only when administered intravenously but also orally. Inactiv-
ity of 1 was attributed to its weak acidity due to intramolecular
hydrogen bond formation.9
Chromones are a class of naturally occurring compounds that
show interesting biological and pharmacological activities coupled
with low toxicity.1a,b
In spite of possessing similar steric demand, fluorine and hydro-
gen analogues differ significantly in their chemical and biological
properties. Fluorinated analogues are generally poorer substrates
for metabolic enzymes, as compared to the hydrogen analogues.
Moreover -F at 6-position in the quinolone structure has invariably
Chromones having heterocyclic substituents at 2 and 3 posi-
tions have been reported to possess antiallergic activity2 muscular
relaxation effect and antimicrobial activity.3 On the other hand,
flavones and their derivatives are commonly used as precursors
for many pharmaceutical products such as anticancer agents.4 It
is also well recognized that incorporation of tetrazole into the
chromone skeleton can significantly enhance the antiallergic
activity.5
* Corresponding author. Tel.: +91 240 2403311; fax: +91 240 2403335.
E-mail address: gillch50@rediffmail.com (C.H. Gill).
Figure 1. Chromones with Carboxylic acid and 1H-tetrazole.
0960-894X/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2008.07.007

S. D. Diwakar et al. / Bioorg. Med. Chem. Lett. 18 (2008) 4678–4681
4679
been associated with improved potency resulting in to the devel-
opment and clinical usage of a range of fluoroquinolones for bacte-
rial infections. It is widely accepted that one or just a few atoms in
an organic molecule can dramatically alter its chemical and biolog-
ical nature, including its potency, stability, lipophilicity and bio-
availability. Thus fluorinated analogues are desired options for
derivatization with the objective of arriving at a potent and phar-
macologically acceptable new chemical entity. Various chemother-
apeutic agents containing fluorine moiety are available for clinical
use,10 and some of them are depicted in Figure 2.
Scheme 1. Synthesis of 5-(4-nitrobenzyl)-1H-tetrazole 5. Reagents and conditions:
(a) Conc. H2SO4/HNO3, 0 °C, 58%; (b) NaN3, ZnBr2, water, reflux, 87%.
The presence of tetrazole and other active groups like chromone
in a single molecule has rarely been divulged. Owing to the versa-
tile bioactivities exhibited by chromones, tetrazole and potential of
fluorine, we directed our efforts to generate libraries of diverse chr-
omones. In continuation of our work on bioactive chromones, we
now wish to disclose our results towards the synthesis and in-vitro
antibacterial assessment of substituted 3-((Z)-2-(4-nitrophenyl)-2-
(1H-tetrazol-5-yl)-vinyl)-4H-chromen-4-ones
strategies.
by
convergent
Scheme 2. Convergent approach. Reagent and condition: (a) Dry pyridine, reflux,
40–88%.
Condensation reactions of 3-formyl chromones with com-
pounds containing active methylene group have been extensively
studied.11–14 Such kind of condensation reactions are achieved by
using either acid or base catalyst. Similarly, the condensation reac-
tions of 3-formyl chromones with hydrazine, mono substituted
hydrazine, hydroxylamine and guanidine have also been stud-
ied.15–17
Based on such reports, we designed linear as well as convergent
route to synthesize 7a–h. In convergent fashion, p-nitrobenzylcya-
nide 418 obtained by nitration of benzyl cyanide was converted
into corresponding tetrazole derivative 519 in 87% yield, by reflux-
ing a mixture of 4, zinc bromide and sodium azide in water for 24 h
(Sharpless method)20–22 Scheme 1. The compound 4 was then con-
densed swiftly with 3-formylchromones 6a–h23 in pyridine24 un-
der reflux to afford title compounds 7a–h25 selectively as a (Z)
regioisomer in 40–88% yields Scheme 2. The (Z) regiochemistry
of the double bond is confiremed by NOE experiment.26
However in linear fashion, the condensation of p-nitro-benzyl
cyanide 4 was carried out with 3-formylchrom-ones 6a in the pres-
ence of sodium acetate in acetic anhydride at 80 °C to give com-
pound 8a27 in 81% yield. Conversion of the cyano group of 8a
into the tetrazole moiety 7a was attempted under different condi-
tions,28a–e but surprisingly the reactions failed in all the tested con-
ditions Scheme 3.
Scheme 3. Linear approach. Reagent and conditions: (a) Sodium acetate, acetic
anhydride, 80 °C, 81%; (b) NaN3, ZnBr2, water, reflux.
(ATCC 13709 ‘Smith’), S. aureus 032, E. faecalis (ATCC 29212) and S.
pneumoniae (ATCC 49619). S. aureus 032 is an methicillin-resistant
S. aureus (MRSA) strain and was obtained as a clinical isolate,29 and
remaining strains were obtained from ATCC, USA. S. aureus (ATCC
13709 ‘Smith’) is a wild type methicillin sensitive strain. Reference
strains used as quality control for MIC testing included S. aureus
ATCC 29213. Erythromycin, Gentamicin, Ampicillin and Ciproflox-
acin were recovered from their commercial preparations. The puri-
ties and potencies of the agents recovered from commercial
preparations were documented by ascertaining the purity of
>98.5% by high-pressure liquid chromatographic analysis and by
showing that the MICs of antibacterials were within acceptable
limits against quality control strains.
All the tetrazolyl chromones 7a–h were assayed for their in-vi-
tro antibacterial activity by MIC determination against a panel of
pathogenic bacterial strains such as E. coli (ATCC 25922), S. aureus
MICs were determined as per CLSI (Clinical and Laboratory
Standards Institute) recommendations on Mueller Hinton Agar
containing serial twofold dilutions of drugs.30 For each strain, ap-
prox. 104 CFU were applied per spot using a multipoint inoculator
(Applied Quality Services, UK). Incubations were done at 35 °C, and
growth was scored at 24 h. In MIC studies twofold differences were
confirmed by a third repetition and more frequent results were re-
ported. MICs employing S. pneumoniae were determined as per
CLSI recommendations on Mueller Hinton Agar supplemented with
yeast extract (0.25%, Difco, USA), glucose (0.5%, Sigma) and sheep
blood (5%). The incubation was carried out in a CO2 incubator
(5% CO2).30 Dimethyl sulfoxide and potent antibacterial drugs
Erythromycin, Ampicillin, Gentamicin and Ciprofloxacin were used
as solvent control and standards, respectively. The results are fur-
nished in Table 1.
Figure 2. Fluorinated drugs.

4680
S. D. Diwakar et al. / Bioorg. Med. Chem. Lett. 18 (2008) 4678–4681
Table 1
Antibacterial activities of substituted 3-((Z)-2-(4-nitrophenyl)-2-(1H-tetrazol-5-yl) vinyl)-4H-chromen-4-ones
Antibacterial activity (MIC,
Microorganism
l )
g mlÀ1
Entry
R1
R2
R3
R4
S. aureusa
S. aureusb
E. faecalis
S. pneumoniae
E. coli
7a
7b
7c
7d
7e
7f
H
H
H
H
H
H
H
H
H
Cl
Br
F
Cl
Cl
CH3
CH3
H
H
H
H
H
CH3
H
H
H
H
H
Cl
H
H
>32
>32
8.0
16
16
08
>32
>32
0.5
2.0
4.0
>32
2.0
>32
>32
4.0
8.0
16
>32
>32
0.5
1.0
16
>32
>32
>32
>32
>32
>32
>32
7g
7h
>32
16
>32
16
H
CH3
8.0
Erythromycin
Ampicillin
Gentamicin
Ciprofloxacin
0.5
>32
>32
0.5
2.0
0.5
>32
0.5
0.12
0.06
>32
>32
>32
>32
>32
0.25
0.25
0.12
>32
2.0
a
S. aureus (ATCC 13709 ‘Smith’).
S. aureus 032 (Is an MRSA strain and was obtained as a clinical isolate).
b
The results indicate that among the tetrazolyl chromone ana-
to the activity of Gentamicin. Erythromycin, Ampicillin and Cipro-
floxacin were found to be inactive against this methicillin-resistant
S. aureus strain indicating the presence of underlying quinolone as
well as macrolide resistance.
logues, 7a, which is unsubstituted at R2 of the chromones ring,
lacks significant activity. Introduction of –Cl, –Br and –CH3 at R2
of chromones ring, thereby producing analogues 7b, 7c and 7g,
respectively, did not alter the activity profile and continued to
demonstrate the lack of activity similar to 7a against the bacterial
strains tested. However, 7g demonstrated a modest indication of
activity against methicillin sensitive strain of S. aureus (S. aureus ).
Amongst the halogens, –F at R2 shows distinct enhancement in
the activity against all the bacterial strains with the exception of E.
coli. Thus –F substituted compound, 7d, exhibited significant drop
As discussed above, with the exception of Gentamicin, other
antibacterial agents studied here such as Erythromycin, Ampicillin
and Ciprofloxacin did not show appreciable anti-MRSA activity. To
date only Vancomycin, Linezolid, Tigecycline and Daptomycin pos-
sess indications for the treatment of MRSA infections. Ciprofloxacin
is a broad-spectrum quinolone antibacterial used commonly for
the treatment of infections caused by Gram-negative pathogens.
Erythromycin, Ampicillin and Gentamicin are primarily used to
treat Respiratory Tract Infections (RTI) infections caused by
Gram-positive pathogens and certain atypical Gram-negative
organisms.
Exploratory studies aimed at understanding the mechanism of
action of 7f, a representative compound described in this work,
were performed employing genetically defined resistant mutants.
Deleterious effect on the activity of 7f against bacterial pathogens
with decreased protein biosynthesis (brought about by addition of
Chloramphenicol to the test medium) indicated a possible ‘protein
biosynthesis inhibition’ mediated mechanism of action. However,
the antibacterial activity of 7f was found to be unaffected against
a range of linezolid-resistant mutants of S. aureus. This result indi-
cates that mechanism of action of 7f is dissimilar to Oxazolidinon-
es. Interestingly, activity of 7f was compromised against
macrolide-resistant strains suggesting a similarity of action be-
tween 7f and the macrolides. Macrolides and Oxazolidinones such
as erythromycin and linezolid, respectively, have been shown to
demonstrate their activity by binding to Domain V of 23S rRNA
and thereby inhibiting the protein biosynthesis process. Since Mac-
rolides do not possess anti-MRSA activity,32 it was very interesting
to note that in spite of mechanism of action-based similarity with
Macrolides, 7f demonstrated a strong evidence of anti-MRSA activ-
ity. Amongst protein biosynthesis inhibitors only Linezolid and
Tigecycline have been described to possess potent activity against
MRSA.33
in MICs (0.5 l
g mlÀ1) against S. aureusà and S. pneumoniae strains.
As a result enhanced potency was observed with 7d as compared
to non-fluorinated analogues. This finding is in agreement with
earlier report, wherein 6-fluorine substitution in Norfloxacin was
shown to improve the gyrase inhibition up to 15- to 18-fold and
cell penetration by another 3- to 4-fold, leading to 10- to 100-fold
decrease in the MIC over its 6-hydrogen derivative.31
Introduction of –Cl at R4 of chromones ring, thereby producing
7e, exhibited enhanced activity as compared to its mono-chloro
analogue 7b. Thus 7e showed a further enhancement in the activity
by at least onefold against S. aureus , twofold against E. faecalis,
fourfold against S. aureusà and fivefold against S. pneumoniae as
compared to 7b.
The considerable improvement in the activity was observed
with 7f, having –CH3 substitution at R3 of chromones ring. This
chemical modification resulted in an increase in activity by at least
onefold against S. aureus, E. faecalis and S. pneumoniae, respec-
tively, and threefold against S. aureusà, as compared to 7b an
unsubstituted R3 analogue.
Similarly, augmentation in the activity was also noticed after
introduction of –CH3 at R4 of chromones ring, thereby producing
7h, a dimethyl analogue of 7g. As compared to 7g, single –CH3 sub-
stitution resulted in an increase in activity by at least onefold
against E. faecalis and S. pneumoniae and two to fourfold against
S. aureus strains, respectively.
Amongst all the compounds synthesized 7d, 7e, 7f and 7h
showed the potential to demonstrate anti-MRSA activity. Of partic-
ular interest was the compound 7d, which showed remarkable
activity against the multi-drug-resistant MRSA strain, comparable
Moreover unlike other anti-MRSA agents such as Vancomycin
and Daptomycin that act by complexing with cell wall, activity of
7f was not affected in the presence of Gram-positive cell wall

S. D. Diwakar et al. / Bioorg. Med. Chem. Lett. 18 (2008) 4678–4681
4681
dry pyridine (2 ml) was refluxed in 15 ml round-bottomed flask for 24 h with
stirring. The reaction mixture was cooled to room temperature and slowly
poured into 20 ml of 2 N HCl. The pale-yellow product which then precipitated
out was filtered and air dried to compound 7.
lysate in the test medium suggesting a lack of ‘cell wall-complex-
ing’ mediated antibacterial action.
In summary, convergent synthesis of substituted 3-((Z)-2-(4-
nitrophenyl)-2-(1H-tetrazol-5-yl)vinyl)-4H-chrom-en-4-ones, 7a–
h were accomplished by combination of 3-formyl chromones 6
and 5-(4-nitrobenzyl)-1H-tetrazole 5. However, most of these tar-
geted chemical entities exhibited modest spectrum of antibacterial
activities against pathogenic bacterial strains. Nevertheless,
compounds 7d, 7e, 7f and 7h displayed significantly improved
anti-MRSA activity providing a very noteworthy lead towards the
optimization of the series described in this paper, with a potential
to result into a broad spectrum anti-MRSA agent.
In conclusion, this study revealed that a range of diverse sub-
stituents on chromones ring, at position R2, R3 and R4 modulate
the antibacterial activity. The study also resulted in diverse com-
pounds, particularly active against MRSA strains, which could
prove useful for further derivatization with the objective to design
highly active compounds against difficult-to-treat multi-drug-
resistant bacterial pathogens.
3-((Z)-2-(4-nitrophenyl)-2-(1H-tetrazol-5-yl)vinyl)-4H-chromen-4-one 7a: Yield:
77%; mp: 248–250 °C dec.; IR (KBr): 3110, 1630, 1570, 1518, 1480, 1353, 1245,
1204, 854, 762 cmÀ1 1H NMR (400 MHz, DMSO-d6): d 6.99–7.05 (m, 2H,
;
ArH), 7.52–7.57 (dt, 1H, ArH), 7.60–7.62 (dd, 1H, ArH, J = 1.6 and 7.6 Hz), 8.42–
8.452 (dd, 2H, ArH, J = 2 and 9.2 Hz), 8.45 (s, 1H, CH), 8.50–8.53 (dd, 2H, ArH,
J = 2 and 9.2 Hz), 9.68 (s, 1H, CH,), 10.62 (s, 1H, NH,); Anal. For C18H11N5O4: C
59.84, H 3.07, and N 19.38. Found: C 59.97, H 3.21, and N 19.51; ESI(Àve):
360.3.
6-Chloro-3-((Z)-2-(4-nitrophenyl)-2-(1H-tetrazol-5-yl) vinyl)-4H-chromen-4-one
7b: Yield: 88%; mp: 213–215 °C dec.; IR (KBr): 3085, 1632, 1517, 1467, 1421,
1351, 1155, 1076, 853, 756 cmÀ1 1H NMR (400 MHz, DMSO-d6): d 7.03–7.05
;
(d, 1H, ArH, J = 9.2 Hz), 7.54–7.58 (m, 2H, ArH), 8.43–8.45 (d, 2H, ArH,
J = 8.4 Hz), 8.46 (s, 1H, CH), 8.51–8.53 (d, 2H, ArH, J = 8.4 Hz), 9.75 (s, 1H, CH,),
10.7 (s, 1H, NH); Anal. For C18H10ClN5O4: C 54.63, H 2.55, and N 17.70. Found: C
54.69, H 2.57, and N 17.64; ESI(Àve): 394.2.
6-Bromo-3-((Z)-2-(4-nitrophenyl)-2-(1H-tetrazol-5-yl) vinyl)-4H-chromen-4-one
7c: Yield: 85%; mp: 230–232 °C dec.; IR (KBr): 3085, 1630, 1517, 1465, 1351,
1278, 1076, 853, 793 cmÀ1 1 H NMR(400 MHz, DMSO-d6): d 6.97–6.99 (d, 1H,
;
ArH, J = 8.0 Hz), 7.65 (s, 1H, ArH), 7.68 (s, 1H, ArH), 8.43–8.45 (d, 2H, ArH,
J = 8.0 Hz), 8.46 (s, 1H, CH), 8.51–8.53 (d, 2H, ArH, J = 8.0 Hz), 9.75 (s, 1H, CH,),
10.73 (s, 1H, NH); Anal. For C18H10BrN5O4: C 49.11, H 2.29, and N 15.91. Found:
49.34, H 2.40, and N 15.98; ESI(Àve): 440.2.
6-Fluoro-3-((Z)-2-(4-nitrophenyl)-2-(1H-tetrazol-5-yl) vinyl)-4H-chromen-4-one
7d: Yield: 80%; mp: 238–239 °C dec.; I.R.: 3110, 1620, 1518, 1479, 1421, 1351,
Acknowledgments
1215, 1194, 854, 759 cmÀ1 1H NMR(400 MHz, DMSO-d6): d 7.02–7.05 (m, 1 H,
;
ArH), 7.38–7.43 (m, 2H, ArH), 8.43–8.45 (dd, 2H, ArH, J = 2 and 7.6 Hz), 8.46 (s,
1H, CH), 8.51–8.53 (dd, 2H, ArH, J = 2 and 7.6 Hz), 9.74 (s, 1H, CH,), 10.45 (s, 1H,
NH); Anal. For C18H10FN5O4: C 57.00, H 2.66, and N 18.46. Found: C 57.21, H
2.71, and N 18.38; ESI(Àve): 378.3.
We greatly acknowledge the Director, Wockhardt Research
Centre, Aurangabad, India, for Antibacterial screening of synthe-
sized compounds. Authors are thankful to the Head, Department
of Chemistry, Dr. BAMU, Aurangabad for constant encouragement
and providing necessary facilities.
6,8-Dichloro-3-((Z)-2-(4-nitrophenyl)-2-(1H-tetrazol-5-yl) vinyl)-4H-chromen-4-
one 7e: Yield: 81%; mp: 233–235 °C dec.; IR (KBr): 3078, 1629, 1518, 1424,
1347, 1319, 1284, 1230, 857, 727 cmÀ1 1H NMR(400 MHz, DMSO-d6): d 7.61–
;
7.62 (d, 1H, ArH, , J = 2.4 Hz), 7.87–7.88 (d, 1H, ArH, J = 2.4 Hz), 8.43–8.45 (d,
2H, ArH, J = 8.4 Hz), 8.5 (s, 1H, CH), 8.52–8.54 (d, 2 H, ArH, J = 8.4 Hz), 9.85 (s,
1H, CH,), 10.77 (s, 1H, NH); Anal. For C18H9Cl2N5O4: C 50.25, H 2.11, and N
16.28. Found: C 50.57, H 2.23, and N 16.38; ESI(Àve): 428.2.
References
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6-Chloro-7-methyl-3-((Z)-2-(4-nitrophenyl)-2-(1H-tetrazol-5-yl) vinyl)-4H-chro-
men-4-one 7f: Yield: 78%; mp: 245–246 °C dec.; IR (KBr): 3070, 1634, 1517,
1470, 1342, 1321, 857, 722 cmÀ1 1H NMR(400 MHz, DMSO-d6): 2.35 (s, 3H,
;
CH3), 7.0 (s, 1H, ArH), 7.61 (s, 1H, ArH), 8.42–8.44 (d, 2H, ArH, J = 9.2 Hz), 8.45
(s, 1H, CH), 8.51–8.53 (d, 2H, ArH, J = 9.2 Hz), 9.95 (s, 1H, CH,), 10.7 (s, 1H, NH);
Anal. For C19H12ClN5O4: C 55.69, H 2.95, and N 17.09. Found: C 55.91, H 3.01,
and N 17.23; ESI(Àve): 408.2.
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6-Methyl-3-((Z)-2-(4-nitrophenyl)-2-(1H-tetrazol-5-yl) vinyl)-4H-chromen-4-one
7g: Yield: 40%; mp: 246–247 °C dec.; 1H NMR(400 MHz, DMSO-d6): d 2.3 (s, 3H,
CH3), 6.93–6.95 (d, 1H, ArH, J = 8 Hz), 7.37–7.4 (dd, 1H, ArH, J = 2 and 8.2 Hz),
7.42 (d, 1H, ArH, J = 2.2 Hz), 8.43–8.46 (d, 2H, ArH, J = 8.0 Hz), 8.5 (s, 1H, CH),
8.53–8.56 (d, 2H, ArH, J = 8.0 Hz), 9.67 (s, 1H, CH,), 10.4 (s, 1H, NH); Anal. For
C19H13N5O4: C 60.80, H 3.49, and N 18.66. Found: C 60.97, H 3.57, and N 18.85;
ESI(Àve): 374.25.
6,8-Dimethyl-3-((Z)-2-(4-nitrophenyl)-2-(1H-tetrazol-5-yl) vinyl)-4H-chromen-
4-one 7h: Yield: 69%; mp: 242–243 °C dec.; 1H NMR(400 MHz, DMSO-d6): d
2.21 (s, 3H, CH3),2.24 (s, 3H, CH3), 7.38 (s, 1H, ArH), 7.40 (s, 1H, ArH), 8.43–8.46
(dd, 2H, ArH, J = 2.4 and 9.2 Hz),8.47 (d, 1H, CH, J = 1.6 Hz), 8.53–8.55 (dd, 2H,
ArH, J = 2.4 and 9.2 Hz), 9.72-9.726 (d, 1H, CH, J = 1.6 Hz), 11.1 (s, 1H, NH);
Anal. For C20H15N5O4: C 61.69, H 3.88, and N 17.99. Found: C 61.84, H 3.91, and
N 18.04; ESI(Àve): 388.3.
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water (100 ml.) was refluxed in 250 ml. round-bottomed flask for 24 h with
vigorous stirring. After completion of reaction, the reaction mixture was cooled
to room temperature, the pH was adjusted to 1.0 with conc. HCl, and the
reaction was stirred for 30 min. to break the complex (p-NO2PhCH2CN)2Zn. The
white solid was filtered, washed with 15 ml  2 1 N HCl and dried at 90 °C to
give 11 g of 2 as a white powder. Yield 87%; mp: 189–190 °C; 1H NMR
(400 MHz, DMSO-d6): 4.37 (s, 2H, CH2), 7.50 (d, 2H, ArH, J = 8.0 Hz), 8.16 (d, 2H,
ArH, J = 8.0 Hz); ESI(Àve): 204.5.
26. NOE experiment is performed on 7b. Irradiation of the methylene proton at C-2
of the chromone ring (d = 9.75) showed no effect on other protons, whereas
irradiation of the vinyl proton (d = 8.46) showed an increase of aromatic
protons (d = 8.43À8.84). Thus, structure as well as regiochemistry (Z) of the
double bond is confirmed.
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