Design, synthesis, and biological evaluation of new cinnamic derivatives as antituberculosis agents

Article abstract of DOI:10.1021/jm101510d

Tuberculosis, HIV coinfection with TB, emergence of multidrug-resistant TB, and extensively drug-resistant TB are the major causes of death from infectious diseases worldwide. Because no new drug has been introduced in the last several decades, new classes of molecules as anti-TB drugs are urgently needed. Herein, we report the synthesis and structure-activity relationships of a series of thioester, amide, hydrazide, and triazolophthalazine derivatives of 4-alkoxy cinnamic acid. Many compounds exhibited submicromolar minimum inhibitory concentrations against Mycobacterium tuberculosis strain (H₃₇Rv). Interestingly, compound 13e, a 4-isopentenyloxycinnamyl triazolophthalazine derivative, was found to be 100-1800 times more active than isoniazid (INH) when tested for its ability to inhibit the growth of INH-resistant M. tuberculosis strains. The results also revealed that 13e does not interfere with mycolic acid biosynthesis, thereby pointing to a different mode of action and representing an attractive lead compound for the development of new anti-TB agents.

Full text of DOI:10.1021/jm101510d

ARTICLE
pubs.acs.org/jmc
Design, Synthesis, and Biological Evaluation of New Cinnamic
Derivatives as Antituberculosis Agents
Prithwiraj De,†,‡ Georges Koumba Yoya,†,‡ Patricia Constant,†,§ Florence Bedos-Belval,†,‡ Hubert Duran,†,‡
||
Nathalie Saffon,†, Mamadou Daffꢀe,*,†,§ and Michel Baltas*,†,‡
†Universitꢀe de Toulouse, UPS, 118, Route de Narbonne, F-31062 Toulouse Cedex 9, France
‡CNRS, LSPCMIB (Laboratoire de Synthꢁese et Physico-Chimie de Molꢀecules d’Intꢀer^et Biologique), 118, Route de Narbonne, F-31062
Toulouse Cedex 9, France
§CNRS, IPBS, (Institut de Pharmacologie et Biologie Structurale), Dꢀepartement Mꢀecanismes Molꢀeculaires des Infections
Mycobactꢀeriennes, 205 route de Narbonne, F-31077 Toulouse Cedex 04, France
Structure Fꢀedꢀerative Toulousaine en Chimie Molꢀeculaire, FR 2599, 118, Route de Narbonne, F-31062 Toulouse Cedex 9, France
S Supporting Information
b
ABSTRACT: Tuberculosis, HIV coinfection with TB, emergence of
multidrug-resistant TB, and extensively drug-resistant TB are the
major causes of death from infectious diseases worldwide. Because no
new drug has been introduced in the last several decades, new classes
of molecules as anti-TB drugs are urgently needed. Herein, we report
the synthesis and structure-activity relationships of a series of thio-
ester, amide, hydrazide, and triazolophthalazine derivatives of 4-
alkoxy cinnamic acid. Many compounds exhibited submicromolar
minimum inhibitory concentrations against Mycobacterium tuberculosis strain (H37Rv). Interestingly, compound 13e, a 4-iso-
pentenyloxycinnamyl triazolophthalazine derivative, was found to be 100-1800 times more active than isoniazid (INH) when
tested for its ability to inhibit the growth of INH-resistant M. tuberculosis strains. The results also revealed that 13e does not interfere
with mycolic acid biosynthesis, thereby pointing to a different mode of action and representing an attractive lead compound for the
development of new anti-TB agents.
’ INTRODUCTION
often a fact.6d Therefore, the urgency and the growing need for the
new class of chemical compounds are well accepted.
Tuberculosis (TB) is a threat to worldwide public health. The
high susceptibility of human immunodeficiency virus (HIV)-
infected persons to the disease1 and the emergence of multidrug-
resistant (MDR) strains2a-2c have brought this infectious disease
into the focus of scientific interest. This fact forced the scientific
community to develop new antimycobacterial agents to treat Myco-
bacterium tuberculosis strains resistant to existing drugs and to shorten
the duration of treatment to improve patient compliance. The lack of
treatment observance is possibly the biggest cause of the occurrence
and spread of MDR strains of M. tuberculosis.3a,3b However, after many
trials,3b,4 two regimens have emerged that involve two periods of
treatment: first, a 2 month long treatment with a set of four drugs,
mainly involving inhibitors of cell envelope component biosynthesis
and particularly fatty acid biosynthesis inhibitors like isoniazid (INH).
This is often followed by 4 months treatment of INH and rifampin.
Hepatotoxicity, being a major side effect in some cases, forces a
premature treatment termination.5 Fluoroquinolones, considered to
be good inhibitors of DNA-based processes, are still in the process of
being established as a second line of anti-TB drugs.6a Studies for the
treatment of MDR TB using the first antibiotic of the oxazolidinone
class, linezolide,6b revealed some cases of peripheral neuropathy as a
side effect.6c Although a number of classes of compounds have been
reported6a with an effect on M. tuberculosis, treatment failure is too
In recent years, trans-cinnamic acid derivatives have attracted
much attention due to their antioxidative,7a antitumor,7b and anti-
microbial7c,7d properties. Piplartin (1),7b an alkaloid and a trans-
cinnamic acid derivative, isolated from Piper tuberculatum, has
been shown to have antitumor as well as antiproliferative activities.
Recently, the synergistic activity8 of trans-cinnamic acid in drug
combinations with INH, rifamycin, and other known antimicrobial
agents against M. tuberculosis has been exemplified. Importantly,
superior intracellular and in vivo activity of a cinnamyl-rifamycin
derivative (2) (Chart 1), in comparison with rifamycin,9 was ob-
served when tested against 20susceptible and MDR M. tuberculosis
strains. Significantly, trans-cinnamic acid was used to treat TB even
before antimicrobial chemotherapy was used.10
We have reported in a preliminary communication11 the syn-
thesis and biological evaluation of various cinnamic thioesters
and amides as potential enoyl-acyl carrier protein (enoyl-ACP)
analogues. Importantly, (E)-N-(2-acetamidoethyl)-3-{4-[(E)-
3,7-dimethylocta-2,6-dienyloxy]phenyl}propanamide (8f) was
found to have an excellent in vitro activity (MIC = 0.1 μg/mL)
Received: November 23, 2010
Published: February 10, 2011
r
2011 American Chemical Society
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|

Journal of Medicinal Chemistry
ARTICLE
Chart 1. Piplartin (1) and Cinnamyl-Rifamycin Derivative
(2)
Chart 2. Illustration of Drug Design Strategy
(EDC HCl) in the presence of 4-N,N-dimethylaminopyridine
3
(DMAP) in dry dichloromethane (Scheme 2) to furnish acet-
amidoethyl 3-(4-alkoxyphenyl)prop-2-enethioate derivatives 7a,
e,f (67-78% yields). N-Acetylethylenediamine, a nitrogen var-
iant of N-acetylcysteamine, was used as a coupling partner of the
enoyl-acids 6a,e,f. The (E)-N-(2-acetamidoethyl)-3-(4-alkoxy-
phenyl) prop-2-enamide derivatives 8a,e,f were prepared in good
yields (63-83%) under similar conditions as described for the
synthesis of 7a,e,f. We synthesized (E)-3-(4-alkoxyphenyl)-N-
(pyridin-2-yl)prop-2-enamide derivatives 9a,e,f (40-48% yields)
using 2-aminopyridine, an aromatic amine as coupling partner, in
against M. tuberculosis. In continuation of our ongoing research
program, directed toward design and synthesis of cinnamoyl
derivatives as anti-TB agents, we describe in this report the syn-
thesis and biological evaluation of new and very potent drug
candidates, an extended family of cinnamyl-thioesters, amides,
hydrazides, and triazolo-phthalazides.
’ CHEMISTRY
To design new cinnamic acid-based drug candidates, we divid-
ed the projected molecules in three parts: the cinnamic acid part
as enoyl-acyl backbone, 4-alkoxy substitution as lipophilicity con-
trol, following Lipinski's12 rules, and attachment of druglike
molecules to the acid functionality (Chart 2). Variations were
accomplished with the choice of alkyl groups in alkoxy substitu-
tion and druglike molecules. Some cyclopropyl, isosteric to the
double bond, derivatives that have been already synthesized13
replacing the enoyl-acyl part were also evaluated to explore the
possible importance of the double bond.
the presence of EDC HCl and DMAP in dichloromethane.
3
In view of the recent report on the cinnamyl-rifamycin deri-
vative,9 we decided to synthesize (2E)-3-(4-methoxyphenyl)-N-
(3-oxo-1,2-oxazolidin-4-yl)prop-2-enamide (10a). To overcome
the poor solubility of D-cycloserine, BSA (bis-trimethylsilylace-
tamide) was used in the coupling reaction between acid chloride,
made from acid (6a), oxalyl chloride, and D-cycloserine in the
presence of N-methylpyrrolidine in dichloromethane to afford
amide 10a in good yield (71%).16 In spite of the fact that D-cyclo-
serine was found to inhibit6a D-alanine racemase and D-alanine D-
alanine ligase enzymes involved in bacterial cell wall biosynthesis,
10a showed anextremelypoorMIC (see biological results) against
M. tuberculosis; thus, no further alkyl modification was pursued.
In continuation of our ongoing effort, we decided to explore
the biological features of 4-alkoxycinnamyl hydrazides as attrac-
tive antituberculosis agents. INH, a frontline drug of TB and an
inhibitor of mycolic acid biosynthesis, was chosen as the coupling
partner of cinnamic acids. Coupling of acids 6a-f with INH was
carried out using N,N,N0,N0-tetra-methyl-O-(1H-benzotriazol-1-
yl)-uronium hexafluorophosphate (HBTU) in the presence of
diisopropylethylamine, in acetonitrile to afford the respective
(E)-N0-[3-(4-alkoxyphenyl)propenoyl]isonicotinohydrazide de-
rivatives (11a-f) in good yields (61-81%). We would like to
mention that (E)-N0-[3-(4-methoxyphenyl)propenoyl]isonico-
tinohydrazide (11a) was found to be identical in all aspects as
reported by Carvalho et al.7d
(E)-4-Hydroxycinnamic acid (3) and (E)-4-methoxycinnamic
acid (6a) are commercially available. The carboxylic function of
4-hydroxycinnamic acid (3) was protected first to introduce
different alkyl chains on the phenolic functionality and used as a
common precursor of most of the amides, thioesters, hydrazides,
and phthalazides obtained. Methyl-ester of (E)-4-hydroxy cin-
namic acid14 was prepared in excellent yield (95%) by refluxing it
in methanol in the presence of a catalytic amount of concentrated
H2SO4 and 4 Å molecular sieves. (E)-Methyl-4-hydroxycinna-
mate (4) was then subjected to alkylation by refluxing suitable
alkyl halide (isopentenyl bromide, geranyl bromide, or ethyl
iodide) in dry acetone in the presence of anhydrous K2CO3 and
KI (used only in case of alkyl bromides) to give corresponding
phenoxy ethers 5c,e,f in good to excellent yields (70-95%).
Because of the presence of a trifluoromethyl functionality, a simi-
lar reaction with 2,2,2-trifluoroethyl iodide resulted in a poor yield
(38%) of (E)-methyl 4-trifluoroethoxycinnamate (5d). There-
fore, we modified the reaction procedure, and 5d was prepared
from 4 using 2,2,2-trifluoroethyl iodide as the alkylating agent and
NaH as the base in dimethyl sulfoxide (DMSO) in 52% yield. (E)-
Methyl 4-trifluoromethoxycinnamate (5b) was obtained (89%
yield) by Wadworth-Emmons coupling reaction between com-
mercially available 4-trifluoromethoxybenzaldehyde (40) and meth-
yl diethylphosphonoacetate under basic conditions15 (Scheme 1).
The methyl-2(4-alkoxyphenyl)cinnamates 5b-f were then saponi-
fied to the corresponding carboxylic acids 6b-f using K2CO3 in
refluxing aqueous methanol as reported in Scheme 1 (97-99%
yields).
To explore the influence of other hydrazides, 1-hydrazinop-
hthalazine hydrochloride, an antihypertensive drug17a,17b of mo-
derate potency, was coupled with acids 6a-f in the presence of
EDC HCl, N-hydroxybenzotriazole (HOBt), and triethylamine
3
to afford (2E,N0,E)-3-(4-alkoxyphenyl)-N0-[phthalazin-1-(2H)-
ylidiene]acrylohydrazides (12a-f) in good yields as reported in
Scheme 3. In addition to the usual characterization, X-ray of 12e
confirms the structure (Chart 3).
However, in a different experimental condition, coupling of acids
6a-fwith 1-hydrazinophthalazine hydrochloride in acetonitrile under
refluxfor48hinthepresenceofEDC HCl, HOBt, and triethylamine
3
The acids 6a,e,f were coupled with N-acetylcysteamine using
1-ethyl-3-(3-dimethyllaminopropyl)carbodiimide hydrochloride
furnished the corresponding 3-(4-alkoxystyryl)-[1,2,4]triazolo[3,4-
R]phthalazines (13a-f) in good yields (65-90%). The X-ray
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Journal of Medicinal Chemistry
ARTICLE
Scheme 1. Synthesis of Four Alkoxycinnamic Acids
Scheme 2. Synthesis of Thioester and Amide Derivatives
structure of the compound 13a (Chart 4) confirmed the formation of
the triazolophthalazine derivative as well as the styryl backbone of the
entire series of compounds issued by a coupling-intramolecular
cyclization-dehydration sequence. The present reaction conditions
established for the synthesis of triazolo-phthalazine derivatives add to
other different methodologies available in the literature.18a-18d
observed when N-acetylethylenediamine was used as a druglike
molecule. While amides 8a and 8e were found to be less active
than their thioester counterparts, the derivative 8f showed a very
potent activity (0.24 μM) as well as cytotoxicity [IC50 = 28 μM;
selectivity index (SI) = 116] (Table 1). The family of compounds
with 2-aminopyridine, an aromatic amine, as a coupling partner
also showed an increase in activity with an increase in lipophi-
licity as compound 9f was found to have a better MIC (2.7 μM)
in comparison to its methyl (9a; 248 μM) and isopentenyl (9e;
52 μM) analogues. Moreover, cytotoxicity (IC50 = 388 μM) and
SI (144) of 2-aminopyridine derivative 9f were encouraging. The
D-cycloserine derivative (10a) was found to have poor activity
(950 μM) as well as a poor lipophilic factor (<2); thus, no further
derivative was made.
’ RESULTS AND DISCUSSION
All of the synthesized compounds were tested for their ability
to inhibit (minimum inhibitory concentration; MIC) the growth
of M. tuberculosis strain H37Rv in a colorimetric microassay based
on the reduction of MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-
diphenyltetrazolium bromide, Sigma] to formazan by metaboli-
cally active cells.19,20
A significant portion of the compounds was found highly ac-
tive in inhibiting M. tuberculosis at a submicromolar level. Among
compounds with N-acetylcysteamine frame, compound 7f with a
geranyl chain was found to be 150-fold more active (1.5 μM)
than its methyl analogue (7a, 225 μM). A similar trend was
Gratifyingly, the series of INH derivatives (11a-f) were
found to show extremely good activity range (0.3-2.3 μM).
The derivative with a methyl chain (11a) was found to be the best
in the series in terms of its activity (0.3 μM) as well as cytotoxi-
city (IC50 = 168 μM; 50 μg/mL) and SI (560). Importantly, the
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ARTICLE
Scheme 3. Synthesis of 4-Alkoxycinnamic Hydrazides
Chart 3. X-ray Structure of 12e
of the parent compound 12e, showed excellent antitubercular
potency (MIC = 1.4 μM), in comparison with other derivatives
in the series, and more importantly, with good cytotoxicity
(IC50 = 449 μM; 160 μg/mL) and SI (SI = 320).
To explore the importance of the enoyl-acyl backbone, with
an approach to replace the double bond by isosteric cyclopropyl
moiety, we also synthesized13 3-[2-(4-alkoxyphenyl)cyclopro-
pyl]-[1,2,4]triazolo[3,4-R]phthalazine (14a-f; racemates; syn-
thesis described elsewhere13), and their in vitro anti-TB potenti-
ality have been evaluated (Table 2). Significantly, the MIC values
of the compounds (14a-f) were found to be poor as compared
to their olefinic analogues (13a-f). In regard to the difference in
activities between the enoyl and the cyclopropyl series, a plau-
sible explanation could be the respective Michael acceptor ability.
Chew et al.22 have recently showed that cinnamaldehydes can act
as Michael acceptors and inhibit thioredoxin reductase through
nucleophilic addition of glutathione cystine -SH residues. In our
case, from a chemical point of view, the compounds having an
electron-withdrawing group in the para-position of the aromatic
ring should be more active to Michael addition. It should be a
clear structure-activity relationship if this is the possible reason
of their activity, that is, 4-OCF3 derivatives are expected to show
better inhibitory activities as compared to their 4-OCH3 analo-
gues. However, this is not the case as compound 11a has a 4-fold
better activity (0.3 μM) as compared to 11b (1.1 μM), and simi-
larly, 13a (53 μM) exhibits approximately 15-fold better acti-
vity than 13b (702 μM). In view of these results, we can suggest
that the Michael addition is not the mode of action of these
compounds.
Because resistance to the current drugs remains a serious
problem, resulting in the occurrence of MDR, which are resistant
to the frontline INH and rifampicin drugs, and extensively drug-
resistant (XDR), we decided to test our newly synthesized com-
pounds on INH-resistant strains. As mutations in InhA23 and katG
represent the main modes of primary resistance to INH in TB
patients,24 we chose to evaluate the inhibitory activity of our com-
poundsontwo INH-resistant strains. MYC5165is a M. tuberculosis
strain mutated in InhA, whereas 1400 is mutated in katG. Two
representative INH derivatives 11a,e and the most active triazo-
lophthalazine derivative 13e were tested on MYC5165 and 1400.
The inhibitory activities of 11a,e were found to be following
similartrends withvalues inthe samelogrange asthatof INH itself
Chart 4. X-ray Structure of 13a
introduction of ethyl (11c), trifluoromethyl (11b), and/or tri-
fluoroethyl (11d) alkyl chains as 4-alkoxy groups still showed
good potency (1.3, 1.1, and 2.2 μM, respectively) and good lipo-
philicity (ClogP) values as antituberculosis agents. The isopen-
tenyl and geranyl derivatives (11e and 11f, respectively) showed
a similar range of MIC results; 11e was found to have a better
potentiality as a drug candidate in view of its good cytotoxicity
(IC50 = 256 μM; 90 μg/mL, SI = 111) profile. Significantly, TLC
analysis21 of methyl esters derivatives obtained after saponifica-
tion of metabolically labeled M. tuberculosis with [1-14C] acetate
in the presence of synthesized molecules showed that 11e, a re-
presentative of the series (11a-f), inhibits mycolic acid bio-
synthesis (Chart 5).
Concerning the family of 1-phthalazine (12a-f), MIC results
were moderate, but the trend of cytotoxic behavior was not acce-
ptable. Interestingly, the combination of isopentenyl chain as a 4-alkoxy
substituent with triazolophthalazine (13e), a modified enoyl-acyl
system obtained by the intramolecular cyclization-dehydration
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Journal of Medicinal Chemistry
ARTICLE
Table 1. Structure and Activities of Cinnamic Derivatives
compd
R1
R2
H37Rv MIC (μM)
toxicity IC50a (μM) (SI)b
CLogP c
PSAd (Å2)
7a11
7e11
7f11
8a11
8e11
8f11
9a11
9e11
9f11
10a
11a
11b
11c
11d
11e
11f
methyl
N-acetylcysteamine
N-acetylcysteamine
N-acetylcysteamine
N-acetylethylenediamine
N-acetylethylenediamine
N-acetylethylenediamine
2-aminopyridine
2-aminopyridine
2-aminopyridine
D-cycloserine
225
48
197 (0.7)
84 (1)
34 (23)
858 (-)
196 (1)
28 (116)
255 (1)
ND
1.65
3.46
5.75
1.08
2.78
4.81
2.54
4.62
6.64
1.24
1.87
2.98
2.40
2.66
3.57
5.60
2.08
3.19
2.61
2.87
3.78
5.81
3.20
4.31
3.73
4.00
4.91
6.94
-0.66
80.32
80.32
80.32
67.43
67.43
67.43
51.22
51.22
51.22
76.66
80.32
80.32
80.32
80.32
80.32
80.32
76.41
76.41
76.41
76.41
76.41
76.41
52.31
52.31
52.31
52.31
52.31
52.31
50.94
isopentenyl
geranyl
methyl
1.5
>1908
186
0.24
248
52
isopentenyl
geranyl
methyl
isopentenyl
geranyl
methyl
2.7
950
0.3
1.1
1.3
2.2
2.3
1.9
50
388 (144)
ND
methyl
INH
168 (560)
ND
CF3
INH
ethyl
INH
ND
CF3CH2
isopentenyl
geranyl
methyl
INH
ND
INH
256 (111)
43 (22)
437 (8)
ND
INH
12a
12b
12c
12d
12e
12f
hydralazine
CF3
hydralazine
21
ethyl
hydralazine
12
ND
CF3CH2
isopentenyl
geranyl
methyl
hydralazine
20
ND
hydralazine
21
26 (1)
56 (0.8)
695 (13)
ND
hydralazine
72
13a
13b
13c
13d
13e
13f
triazolophthalazine
triazolophthalazine
triazolophthalazine
triazolophthalazine
triazolophthalazine
triazolophthalazine
53
CF3
702
39
ethyl
ND
CF3CH2
isopentenyl
geranyl
170
1.4
19
ND
449 (320)
259 (13)
>3649 (>6081)
INH
0.6
a Fifty percent inhibitory concentration (cytotoxicity toward THP-1 cells). b Selectivity index: ratio of cytotoxicity to in vitro activity against M.
tuberculosis (IC50/Mtb MIC). c CLogP calculated using the ChemDraw Ultra, version 10.0, software by Cambridge Soft. ND, not done. d Polar surface
area (PSA): Calculated using Calculator Plugin Marvin in www.chemaxon.com.
Chart 5. Radio Thin-Layer Chromatography Analysis of
Mycolic Acids from M. tuberculosis Treated with Different
Doses of Selected Compoundsa
Table 2. Anti-TB Activities of Cyclopropyl Derivatives
compd
R1
H37Rv MIC (μM)
CLogP
14a
14b
14c
14d
14e
14f
methyl-
CF3-
395
170
378
1302
21
2.63
3.74
3.16
3.12
4.33
6.36
ethyl
CF3CH2-
isopentenyl-
geranyl-
28
aFor each compound: left, MIC; right, MIC/5.
0.2 μM) and 1800-fold better activity against 1400 strain (13e;
MIC = 0.4 μM) as compared to INH (MIC = 18 and 729 μM,
respectively). The importance of the isopentenyl side chain,
cinnamic double bond, and triazolophthalazine part is evident
from these biological results as none of the other triazolophtha-
lazine derivatives (13a-d,f) are active enough. Furthermore, the
as represented in Table 3, thus not allowing at the moment to
propose these compounds as INH prodrugs or not.
Finally, to our great delight, compound 13e showed 100-fold
better in vitro activity against MYC5165 strain (13e; MIC =
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ARTICLE
Table 3. Anti-TB Activities against INH-Resistant M. tuber-
culosis Strains to INH
Chart 6. Enumeration of Molecules
compd
MYC5165 MIC (μM)
1400 MIC (μM)
11a
16
27
0.2
18
5
320
68
11e
13e
0.4
729
5
INH
ciprofloxacine
Toxicity. Cytotoxicities (IC50; 50% inhibitory concentration) toward
THP-1 cells were determined for some relevant molecules by using the
Graph Pad Prism 5.0 Software after measuring the extent of reduction of
MTT following 72 h of exposure to compounds. Corresponding SIs
have been calculated (IC50/MIC).
radio thin-layer chromatography analysis (Chart 5) revealed that
compound 13e does not inhibit mycolic acid biosynthesis. This
fact, in combination with its inhibitory efficacy against INH-
resistant strains, suggests that compound 13e may be considered
as a good hit, in terms of MIC values, cytotoxicity profile (SI =
320), CLogP (4.91), and PSA (52.31 Å2)25 values with a mode of
action to be discovered.
Evaluation of the in Vivo Effects of Compounds on Mycolic Acid
Biosynthesis21. The synthesized compounds were added at two con-
centrations (MIC and MIC/5) to broth cultures of M. tuberculosis in the
exponential growth phase. After 8 h of incubation, [1-14C]acetate (2 μCi,
13 μM) was added to follow the biosynthesis of lipid products. After 24 h,
bacteria were treated with 40% KOH (w/v) in methoxyethanol (1:7)
overnight at 100 °C. The suspensions were then acidified by addition of
20% H2SO4. Lipids, including mycolic acids, were then extracted with
diethyl ether. The crude fatty acids were methylated, and methyl ester
derivatives were analyzed by thin-layer chromatography on Silica Gel 60
(Macherey-Nagel) run in dichloromethane followed by phosphorimaging
(Variable Mode Imager Typhoon TRIO, Amersham Biosciences).
Chemistry. Organic solvents were purified when necessary by stan-
dard methods26 or were purchased from Aldrich. Melting points (mp)
were obtained on a Buchi apparatus and are uncorrected. Infrared (IR)
spectra were recorded on a Perkin-Elmer 1725 infrared spectrophoto-
meter, and the data are reported in inverse centimeters. Proton nuclear
magnetic resonance (1H NMR) spectra were obtained with a Bruker
AC-300 spectrometer. Chemical shifts were reported in parts per million
(ppm) relative to tetramethylsilane (TMS), and signals are given as
follows: s, singlet; d, doublet; t, triplet; and m, multiplet. For better NMR
data analysis, compounds were numerated as follow (Chart 6). Mass
spectra were recorded on an R 10-10 C Nermag (70 eV) quadripolar
spectrometer using desorption chemical ionization (DCI), electrospray
(ES), or fast atomic bombardment (FAB) techniques. Analytical HPLC
was carried out on Acquity Waters HPLC system with a Acquity BEH
C18 1.7 μm, 21 mm ꢀ 50 mm column and PDA eλ Waters UV detector.
The flow rate was 0.3 mL/min with gradient eluation over 6 min, from
70% CH3CN-H2O to 100% CH3CN with 0.1% TFA. All of the com-
pounds tested for the antitubercular activity were at least 95% pure as
determined by HPLC (Supporting Information).
’ CONCLUSIONS
New cinnamic acid-based molecules were synthesized by
simple, clean, and efficient synthetic protocols. Introduction of
selective alkyl groups as lipophilicity control was achieved suc-
cessfully. The importance of the double bond was also probed.
Many of the synthesized molecules have encouraging antituber-
culosis activities (MIC) and cytotoxicity ranges toward THP-1
cells and possess satisfactory druggability (PSA = 50-80 Å2).
The compounds (E)-N-(pyridin-2-yl)-3-[4-{(E)-3,7-dimethyl-
octa-2,6-dienyloxy}phenyl]prop-2-enamide (9f) and (E)-3-[4-
(3-methylbut-2-enyloxy)styryl]-[1,2,4]triazolo[3,4-R]phthalazine
(13e) were found to have good anti-TB activity, and both are
compatible in terms of cytotoxycity and ClogP values. Moreover,
the entire series of INH derivatives (11a-f) have good anti-TB
activities. Importantly, the MIC value of 11a against M. tuberculosis
H37Rv strain is comparable to, if not even better than that of, INH,
a frontline anti-TB drug. Compound 11e was found to inhibit the
biosynthesis of mycolic acids, as does INH. Further development
of the INH derivatives is underway, in particular, biomimetic and
enzymatic studies, to elucidate their mode of action. Finally and
most importantly, compound 13e does not inhibit mycolic acid
biosynthesis but is extremely active against two INH-resistant
strains. Thus, 13e may be considered as a hit in terms of excellent
antitubercular activity, lipophilicity, and PSA values. The compre-
hension of the mode of action of 13e is one of the goals of the
ongoing research program in our laboratories.
(E)-Methyl-3-(4-hydroxyphenyl) Propenoate (3). Compound 1 was
synthesized by the reported procedure.14
Preparation of (E)-Methyl-3-(4-alkoxyphenyl)prop-2-enoate (5a-
f): General Procedure. To compound 4 (0.5 g, 2.8 mmol, 1 equiv) in dry
acetone (15 mL) was added KI (added only in the case of alkyl bromide;
0.6 g, 4.2 mmol, 1.5 equiv), K2CO3 (0.58 g, 4.2 mmol, 1.5 equiv), and
alkyl halide (4.2 mmol, 1.5 equiv). The reaction mixture was refluxed for
20 h and then cooled to room temperature and filtrated. The filtrate was
evaporated to dryness, and water (20 mL) was added into the residue
and extracted with ethyl acetate (20 mL ꢀ 3). The combined organic
layer was dried with Na2SO4 and evaporated in vacuo. The residue was
purified by chromatography over silica gel (20% ethyl acetate in
petroleum ether) to afford compound 5a-f as white solids.
’ EXPERIMENTAL SECTION
Biology. Inhibition of Mycobacterial Growth. The susceptibility
of M. tuberculosis strain H37Rv to all synthesized compounds was
evaluated by determining the MIC. We used a colorimetric microassay
based on the reduction of MTT (Sigma) to formazan by metabolically
active cells.19,20 Briefly, serial 2-fold dilutions of each drug solubilized in
DMSO were prepared in 7H9 broth [Middlebrook 7H9 broth base
(Difco)] using 96-well microtiter plates, and 100 μL of M. tuberculosis
H37Rv suspension in 7H9 broth was added to each well. After 6 days of
incubation, MTT was added (50 μL, 1 mg/mL). After 1 day of incuba-
tion, solubilization buffer was added to each well. The optical densities
were measured at 570 nm. The MIC was determined as the lowest
concentration of drug that inhibited bacterial growth (absorbance from
untreated bacilli was taken as a control for growth). Reported MICs are
an average of three individual measurements.
(E)-Methyl-3-(4-ethoxyphenyl)prop-2-enoate (5c). Compound 5c
was prepared from ester 4 according to procedure by using ethyl iodide.
Solid (0.40 g, 70%); mp 57-58 °C. IR (KBr) ν cm-1: 2977 (C-H),
1708 (CdO), 1604 (CdC arom.), 1513 (CdC arom.), 1254 (O-C
1
ether). H NMR (CDCl3, 300 MHz) δ ppm: 1.42 (t, 3H, J = 7.0 Hz,
1454
dx.doi.org/10.1021/jm101510d |J. Med. Chem. 2011, 54, 1449–1461

Journal of Medicinal Chemistry
ARTICLE
0
0
H2 ), 3.79 (s, 3H, -COOCH3, H10), 4.06 (q, 2H, J = 7,0 Hz, H1 ), 6.30
(d, 1H, J = 16.0 Hz, H8), 6.90 (d, 2H, J = 8.8 Hz, H3,5), 7.46 (d, 2H, J =
8.8 Hz, H2,6), 7.64 (d, 1H, J = 16.0 Hz, H7). 13C NMR (CDCl3, 75 MHz)
δ ppm: 14.7 (CH3), 51.5 (COOCH3, C10), 63.6 (CH2), 114.7 (C3,5),
115.1 (C8), 126.9 (C1), 129.1 (C2,6), 144.5 (C7), 160.7 (C4), 167.7
(C9). MS (DCI, CH4, pos.) m/z: 207.1 (MHþ).
(E)-Methyl-3-[4-(20,20,20-trifluoroethoxy)phenyl]prop-2-enoate
(5d). A clean dry round-bottom flask (50 mL) with a stir bar was charged
with NaH (60% dispersion in mineral oil, 240 mg, 6 mmol, 1.2 equiv)
under argon. Dry dimethylsulfoxide (4 mL) was added to the reaction
flask and stirred at 0 °C for 15 min. A solution of phenol (4) (890 mg,
5 mmol, 1 equiv) in DMSO (2 mL) was added to the suspension slowly
over 10 min (1 mL of DMSO was used for rinsing). It was then allowed to
stir at 0 °C for 30 min when a dark yellow solution resulted. 2,2,2-
Trifluoroethyl iodide (1.5 mL, 15 mmol, 3 equiv) was added to the
reaction flask. The reaction was then stirred at 80 °C for 24 h and
monitored by TLC (1/9 ethyl acetate/petroleum ether). A saturated
aqueous NH4Cl (2 mL) solution was added to quenchthe reaction. Water
(20 mL) was added and then extracted with diethyl ether (30 mL ꢀ 3).
The organic layer was dried over MgSO4. The solvent was removed in
vacuo, and the residue was purified bysilica gel chromatography (3% ethyl
acetate in petroleum ether) to afford compound 5d as a solid (0.68 g,
52%); mp 96-98 °C. IR (KBr) ν cm-1: 2951 (C-H), 1708 (CdO),
1604 (CdC arom.), 1515 (CdC arom.), 1245 (C-O), 1170 (C-F),
974 (CHdCH), 830 (C arom. 1-4). 1H NMR (CDCl3, 300 MHz) δ
ppm:3.80 (s, 3H, H10), 4.38 (q, 2H, J= 8.0 Hz, CH2), 6.34(d, 1H, J = 16.0
Hz, H8), 6.95 (d, 2H, J = 8.8 Hz, H3,5), 7.50 (d, 2H, J = 8.5 Hz, H2,6), 7.65
(d, 1H, J = 16.0 Hz, H7). 13C NMR (CDCl3, 75 MHz) δ ppm: 51.69
(C10), 65.66 (q, J = 35.9 Hz, CH2), 115.18 (C2,6), 116.51 (C8), 123.15 (q,
(E)-Methyl-3-[4-(3-methylbut-2-enyloxy)phenyl]prop-2-enoate
(5e). Compound 5e was prepared from ester 4 according to the
procedure by using 3,3-dimethylallyl bromide. White solid (0.59 g,
86%); mp 55-57 °C. IR (KBr, νmax) cm-1: 2945 (C-H), 1716
(CdO), 1635 (CdC arom.), 1513 (CdC arom.), 1251(O-C
ether). H NMR (CDCl3, 300 MHz) δ ppm: 1.69 (s, 3H, H4 ),
1
0
0
0
1.74 (s, 3H, H5 ), 3.73 (s, 3H, CH3), 4.49 (d, 2H, J = 6.6 Hz, H1 ),
5.42 (th, 1H, J = 6.6 Hz, J = 1.5 Hz, H2 ), 6.24 (d, 1H, J = 15.9 Hz,
0
H8), 6.84 (d, 2H, J = 8.5 Hz, H3,5), 7.41 (d, 2H, J = 8.5 Hz, H2,6), 7.60
(d, 1H, J = 15.9 Hz, H7). 13C NMR (CDCl3, 75 MHz) δ ppm: 14.20
0
0
0
(C4 ), 39.51 (C5 ), 51.55 (C10), 64 0.91 (C1 ), 114.89 (C3, 5), 115.14
0
0
(C8), 119.22 (C2 ), 126.99 (C1), 129.70 (C2, 6), 138.66 (C3 ), 144.60
(C7), 160.74 (C4), 167.80 (C9). MS (DCI, NH3, pos.) m/z: 264.2
(MNH4þ), 264.2 (MNH4þ).
(E)-Methyl-3-{4-[(E)-3,7-dimethylocta-2,6-dienyloxy]phenyl}prop-
2-enoate (5f). Compound 5f was prepared from ester 4 according to
procedure for 5e synthesis by using geranyl bromide. Solid (0.84 g,
95%); mp 72-75 °C. IR (KBr) ν cm-1: 3168 (CdC-H ethyl.), 2942
(C-H), 2854 (C-H CH3-O), 1720 (CdO), 1637 (CdC ethyl.),
1603 (CdC ethyl.), 1572 (CdC arom.), 1511 (CdC arom.). 1H NMR
0
J = 277.9 Hz, C2 ), 128.90 (C1), 129.61 (C3,5), 143.87 (C7), 158.80 (C4),
167.52 (C9). 19F NMR (CDCl3 282 MHz) δ ppm: -73,85 (t, 3F, J = 8
Hz). MS (DCI, CH4, pos.) m/z: 261.07 (MHþ).
0
0
Representative Procedure: (E)-3-[4-(3-Methylbut-2-enyloxy)phenyl]-
prop-2-enoic Acid (6e). To compound 5e (0.5 g, 2.03 mmol, 1 equiv) in
MeOH (10 mL) was added K2CO3 (1.4 g, 10.1 mmol, 5 equiv) dissolved
in H2O (10 mL). The reaction mixture was refluxed for 3 h, and then,
MeOH was removed under reduced pressure. It was then cooled at 0 °C
and acidified to pH 2 with HCl (1M). The mixture was extracted with
diethyl ether (30 mL ꢀ 3). The combined organic layer was washed with
brine (30 mL), dried with Na2SO4, and evaporated in vacuo to give 6e as
white solid (0.37 g, 91%); mp 148-150 °C. IR (KBr) ν cm-1: 3268 (N-
H), 3168 (CdC-H), 2942 (C-H), 2854 (C-H, CH3-O), 1720
(CDCl3, 300 MHz) δ ppm: 1.60 (s, 3H, CH9 ), 1.67 (s, 3H, H4 ), 1.74 (s,
0
0
0
3H, H10 ), 2.10 (m, 4H, H5 and H6 ), 3.79 (s, 3H, CH3), 4.57 (d, 2H, J =
0
0
0
6.6 Hz, H1 ), 5.07 (m, 1H, H7 ), 5.48 (t, 1H, = 6.5 Hz, H2 ), 6.31 (d, 1H,
J = 15.9 Hz, H8), 6.91 (d, 2H, J = 8.9 Hz, H3,5), 7.47 (d, 2H, J = 8.9 Hz,
H2,6), 7.64 (d, 1H, J = 15.9 Hz, H7). 13C NMR (CDCl3, 75 MHz) δ
ppm: 16.68 (C9 ), 25.67 (C4 ), 26.26 (C6 ), 39.17 (C10 ), 39.51 (C5 ),
51.53 (CH3), 64 0.88 (C1 ), 110.03 (C8), 115.11 (C3, 5), 118.84 (C2 ),
123.73 (C7 ), 126.97 (C1), 129.69 (C2, 6), 131.83 (C8 ), 141.66 (C3 ),
144.61 (C7), 160.16 (C4), 167.78 (C9). MS (APCI, MeOH, pos.) m/z:
315.25 (MHþ).
0
0
0
0
0
0
0
0
0
0
(CdO), 1637 (CdC), 1603 (CdC), 1572 (CdC arom.), 1511
Preparation of (E)-Methyl-3-[4-(trifluoromethoxy)phenyl]prop-2-
enoate (5b). A clean dry round-bottom flask (100 mL) with a stir
bar was charged with NaH (60% dispersion in mineral oil, 950 mg, 23.7
mmol, 1.5 equiv) under argon. It was then washed with petroleum ether
(5 mL ꢀ 2). Dry THF (40 mL) was added to the reaction flask and
stirred at 0 °C for 15 min. Methyldiethylphosphonoacetate (3.5 mL,
19 mmol, 1.2 equiv) was added to the suspension over 10 min. It was then
stirred at 0 °C for 30 min when a colorless solution resulted. A solution of
4-trifluoromethoxybenzaldehyde (2.3 mL, 15.8 mmol, 1 equiv) in dry
tetrahydrofurane (12 mL) was slowly added to the reaction flask over
10 min. The reaction was then stirred at room temperature (25 °C) for
20 h and was monitored by TLC. A saturated aqueous NH4Cl (2 mL)
solution was then added to quench the reaction. Tetrahydrofurane was
removed under reduced pressure, and water (30 mL) was added to the
reactionmixture. Itwas thenextractedwith ethylacetate (60mLꢀ 3). The
organic layer was dried over MgSO4. The solvent was removed in vacuo,
and the residue was purified by silica gel chromatography (3% ethyl
acetate in petroleum ether) to afford compound 5b as a solid (3.3 g, 89%);
mp 48-50 °C. IR (KBr) ν cm -1: 2956 (C-H), 1713 (CdO), 1641
(CdC arom.), 1510 (CdC arom.), 1284 (C-O ether), 1161 (C-F),
1
0
(CdC arom.). H NMR (CDCl3, 300 MHz) δ ppm: 1.69 (s, 3H, H4 ),
0
0
0
1.74 (s, 3H, H5 ), 4.49 (d, 2H, J = 5 Hz, H1 ), 5.42 (t, 1H, = 5 Hz, H2 ),
6.25 (d, 1H, J = 15.6 Hz, H8), 6.86 (d, 2H, J = 8.5 Hz, H3,5), 7.43 (d, 2H, J
= 8.5 Hz, H2,6), 7.67 (d, 1H, J = 15.6 Hz, H7). 13C NMR (CDCl3, 75
0
0
0
MHz) δ ppm: 18.24 (C4 ), 25.83 (C5 ), 64 0.96 (C1 ), 114.20 (C8),
0
0
114.97 (C3, 5), 119.16 (C2 ), 126.69 (C1), 130.11 (C2, 6), 138.75 (C3 ),
146.80 (C7), 161.11 (C4), 172.90 (C9). HRMS (DCI, CH4, pos.) m/z:
calcd for C14H17O3 (MHþ), 233.1178; obtained, 233.1180.
(E)-3-[4-(Trifluoromethoxy)phenyl]prop-2-enoic Acid (6b). Compound
6b was prepared from ester 5b according to procedure used for 6e synthesis.
Solid (0.47 g, quantitative); mp 180-182 °C. IR (KBr) ν cm-1: 3462(OH),
1686 (CdO), 1629 (CdC arom.), 1510 (CdC arom.), 1268 (C-Oether),
1216 (C-F), 931 (CHdCH), 832 (C arom. 1-4). 1H NMR (acetone-d6,
300 MHz) δ ppm: 6.57 (d, 1H, J = 16.1 Hz, H8), 7.39 (d, 2H, J = 8.8 Hz,
H3,5), 7.7 (d, 1H, J = 16.1 Hz, H7), 7.84 (d, 2H, J = 8.6 Hz, H2,6). 13C NMR
(acetone-d6, 75 MHz) δ ppm: 121.83 (C8), 122.59 (q, J = 256.2 Hz, CF3),
123.42 (C3,5), 132.10 (C2,6), 135.94 (C1), 144.89 (C7), 152.20 (C4), 166.78
(C9). 19F NMR (acetone-d6, 282 MHz) δ ppm: -58.52 (s, 3F, CF3O-).
HRMS: calcd for C10H8F3O3 (MHþ), 233.0426; found, 233.0427.
1
(E)-3-(4-Ethoxyphenyl)prop-2-enoic Acid (6c). Compound 6c was
prepared from ester 5c according to procedure used for 6e synthesis.
Solid (0.40 g, quantitative); mp 184-185 °C. IR (KBr) ν cm-1: 3431
(OH), 2980 (C-H), 1679 (CdO), 1600 (CdC arom.), 1510 (CdC
arom.), 1248 (C-O). 1H NMR (CDCl3, 300 MHz) δ ppm: 1.43 (t, 3H,
990 (CHdCH ethylenic), 839 (C arom 1-4). H NMR (300 MHz,
CDCl3) δ: 3.81 (s, 3H, H10), 6.41 (d, 1H, J = 16.0 Hz, H8), 7.23 (d, 2H,
J = 8.7 Hz, H3,5), 7.54 (d, J = 8.9 Hz, H2,6), 7.66 (d, J = 16.0 Hz, H7). 13
C
NMR(75MHz, CDCl3) δ: 51.62 (C10), 118.59 (C8), 120.17 (q, J= 258.1
Hz, CF3), 120.97 (C3,5), 129.29 (C2,6), 132.80 (C1), 142.89 (C7), 150.21
(C4), 166.89 (C9). 19F NMR (CDCl3 282 MHz) δ ppm: -57.79 (s, 3F,
CF3O-). HRMS (APCI, MHþ) calcd for C11H10F3O3, 247.0582; found,
247.0575.
0
0
J = 7.0 Hz, H2 ), 4.8 (q, 2H, J = 7.0 Hz, H1 ), 6.1 (d, 1H, J = 15. Hz, H8),
6.90 (d, 2H, J = 8.7 Hz, H3,5), 7.49 (d, 2H, J = 8.7 Hz, H2,6), 7.74 (d, 1H,
J = 15.9 Hz, H7). 13C NMR (CDCl3, 75 MHz) δ ppm: 14.73 (C2 ), 63.68
0
1455
dx.doi.org/10.1021/jm101510d |J. Med. Chem. 2011, 54, 1449–1461

Journal of Medicinal Chemistry
ARTICLE
0
(C1 ), 114.44 (C8), 114.88 (C3,5), 126.62 (C1), 130.14 (C2,6), 146.84
(C12), 188.0 (C9). EI MS (m/z): 279 (Mþ), 161 (Mþ - Cys), 133
(Mþ - Cys-CO). Anal. calcd for C14H17NO4S: C, 56.93; H, 5.80; N,
5.28. Observed: C, 57.01; H, 5.66; N, 4.79).
(E)-S-2-Acetamidoethyl 3-{4-[(E)-3,7-Dimethylocta-2,6-dienyloxy]-
phenyl}prop-2-enethioate (7f). Compound 7f was prepared from acid
(C7), 161.19 (C4), 172.41 (C9). HRMS (DCI, CH4, pos.) m/z: calcd for
C11H13O3 (MHþ), 193.0865; obtained, 193.0870.
(E)-3-[4-(20,20,20-Trifluoroethoxy)phenyl]prop-2-enoic Acid (6d). Com-
pound 6d was prepared from ester 5d according to procedure used for 6e
synthesis. Solid (0.50 g, quantitative); mp 205-207 °C. IR (KBr) ν cm-1
:
6f according to procedure used for 7e synthesis. Sticky solid (0.54 g,
1
0
3436 (OH), 2921 (C-H), 1683 (CdO), 1603(CdC arom.), 1511 (CdC
67%). H NMR (CDCl3, 300 MHz) δ ppm: 1.53 (s, 3H, H9 ), 1.60 (s,
0
0
0
0
arom.), 1294 (C-O ether), 1174 (C-F), 973 (CHdCH), 830 (C arom.
1-4). H NMR (CDCl3, 300 MHz) δ ppm: 4.57 (q, 2H, J = 8.5 Hz, H1 ),
3H, H4 ), 1.67 (s, 3H, H10 ), 1.91 (s, 3H, C13), 2.03 (m, 4H, H5 ,6 ), 3.08
1
0
(t, 2H, J = 6.4 Hz, CH2S), 3.42 (q, 2H, J = 6.4 Hz, H10), 4.50 (d, 2H, J =
0
0
0
6.37 (d, 1H, J = 16.0 Hz, H8), 7.04 (d, 2H, J = 8.8 Hz, H3,5), 7.57 (d, 2H, J =
8.5 Hz, H2,6), 7.61 (d, 1H, J = 16.0 Hz, H7). 13C NMR (CDCl3, 75 MHz) δ
ppm: 66.45 (q, J = 35.2 Hz, CH2), 116.56 (C3,5), 118.17 (C8), 125.28 (q, J =
277.0 Hz, CF3), 130.21 (C1), 131.28 (C2,6), 145.22 (C7), 160.32 (C4),
168.41 (C9).19FNMR:(acetone-d6, 282 MHz) δppm: -74.65 (t, 3F, J=8.5
Hz, CF3). HRMS: calcd for C11H11F3O3 (MHþ), 247.0583; found,
247.0582.
(E)-3-{4-[(E)-3,7-Dimethylocta-2,6-dienyloxy]phenyl}prop-2-enoic
Acid (6f). Compound 6f was prepared from ester 5f according to pro-
cedure used for 6e synthesis. Solid (0.49 g, 71%); mp 115-117 °C. IR
(KBr) ν cm-1: 3429 (O-H), 2968 (CdCH), 2925 (C-H), 2856 (C-
H), 1671 (CdO), 1626 (CdC), 1602 (CdC), 1573 (CdC arom.),
1512 (CdC arom.), 1246 (C-O ether), 991 (CHdCH trans), 827 (C
arom 1-4). H NMR (CDCl3, 300 MHz) δ ppm: 1.54 (s, 3H, H9 ), 1.61
(s, 3H, H4 ), 1.64 (s, 3H, H10 ), 2.04 (m, 4H, H5 ,6 ), 4.51 (d, 2H, J = 6.9
Hz, H1 ), 5.03 (m, 1H, H7 ), 5.41 (t, 1H, J = 6.9 Hz, H2 ), 6.25 (d, 1H, J =
15.9 Hz, H8), 6.85 (d, 2H, J = 8.7 Hz, H3,5), 7.42 (d, 2H, J = 8.7 Hz, H2,6),
7.66 (d, 1H, J = 15.9 Hz, H7). 13C NMR (CDCl3, 75 MHz) δ ppm: 16.71
6.5 Hz, H1 ), 5.01 (m, 1H, H7 ), 5.40 (tt, J = 6.6 Hz, J = 1.0 Hz, 1H, H2 ),
6.0 (bs, 1H, NH), 6.54 (d, 1H, J = 15.7 Hz, H8), 6.85 (d, 2H, J = 8.7 Hz,
H3,5), 7.42 (d, 2H, J = 8.7 Hz, H2,6), 7.52 (d, 1H, J = 15.7 Hz, H7). 13
C
0
0
0
NMR (CDCl3, 75 MHz) δ ppm: 16.6 (C4 ), 17.6 (C9 ), 23.1 (C10 ), 25.6
0
0
0
(C13), 26.2 (C6 ), 28.4 (C10), 39.4 (C5 ), 39.8 (C11), 65.0 (C1 ), 114.9
0
0
(C3, 5), 118.9 (C2 ), 122.0 (C7 ), 122.2 (C7),126.3 (C1), 130.2 (C2, 6),
0
0
131.8 (C8 ), 141.2 (C8), 141.7 (C3 ), 161.2 (C4), 170.8 (C12), 190.03
(C9). MS (DCI, NH3, pos.) m/z: 402.0 (MHþ), 420.0 (M þ NH4þ).
HPLC purity: 100%.
Representative Procedure: Preparation of (E)-N-(2-Acetamidoethyl)-
3-[4-(3-methylbut-2-enyloxy)phenyl]prop-2-enamide (8e). A clean dry
round-bottom flask (10 mL) with a magnetic stir bar was charged with
carboxylic acid 6e (0.05 g, 0.28 mmol, 1 equiv), N-acetylethylenediamine
(0.043 g, 0.42 mmol, 1.5 equiv), DMAP (0.037 g, 0.31 mmol, 1.1 equiv),
1
0
0
0
0
0
and EDC HCl (0.06 g, 0.31 mmol, 1.1 equiv) under argon. Dry dichloro-
3
methane (4 mL) was added to it and stirred at room temperature for 24 h.
TLC was monitored in dichloromethane/methanol (95/5). Dichloro-
methane was removed under reduced pressure. Ethylacetate (60 mL) was
added to the crude yellow mass, and the solution was thoroughly washed
with water (30 mL ꢀ 3). The organic layer was then dried over MgSO4.
Removal of the solvent gave a crude yellowish mass. It was purified over
silica gel (70-200 mesh) using 10% methanol in ethylacetate to afford 8e
as a white solid (0.055 g, 63%); mp 177-179 °C. IR (KBr) ν cm-1: 3289
(N-H), 3091 (CdC-H), 1654 (CdO), 1620 (CdC), 1605 (CdC
0
0
0
0
0
0
0
0
0
(C4 ), 17.71 (C9 ), 25.68 (C10 ), 26.26 (C6 ), 39.53 (C5 ), 65.05 (C1 ),
0
0
114.47 (C8), 115.11 (C3, 5), 118.97 (C2 ), 123.73 (C7 ), 126.66 (C1),
0
0
130.08 (C2, 6), 131.83 (C8 ), 141.77 (C3 ), 146.79 (C7), 161.13 (C4),
172.32 (C9). MS (DCI, NH3, pos.) m/z: 301 (MHþ), 318 (MNH4þ).
Representative Procedure: Preparation of (E)-S-2-Acetamidoethyl
3-[4-(3-Methylbut-2-enyloxy)phenyl]prop-2-enethioate (7e). A clean
dry round-bottom flask (25 mL) with a magnetic stir bar was charged
with carboxylic acid 6e (0.46 g, 2 mmol, 1 equiv), N-acetylcysteamine
(0.24 g, 2 mmol, 1 equiv), DMAP (0.26 g, 2.2 mmol, 1.1 equiv), and
arom.), 1561,1511 (CdC arom.), 1228 (O-C arom), 972 (CHdCH),
1
0
824 (arom 1-4). H NMR (CDCl3, 300 MHz) δ ppm: 1.76 (s, 3H, H4 ),
0
0
1.81 (s, 3H, H5 ), 2.01(s, 3H, H13), 3.49 (m, 4H, CH2N), 4.54 (d, 2H, H1 ),
0
5.5 (t, 1H, J = H2 ), 6.32 (d, 1H, J = 15.6 Hz, H8), 6.63 (m, 2H, NH), 6.90
EDC HCl (0.42 g, 2.2 mmol, 1.1 equiv) under argon. Dry dichlorome-
3
(d, 2H, J = 8.7 Hz, H3,5), 7.45 (d, 2H, J = 8.7 Hz, H2,6), 7.58 (d, 1H, J = 15.6
thane (15 mL) was added to it and stirred at room temperature for 24 h.
TLC was monitored in ethylacetate. Dichloromethane was removed
under reduced pressure. Ethylacetate (60 mL) was added to the crude
yellow mass, and the solution was thoroughly washed with water (30
mL ꢀ 3). The organic layer was then dried over MgSO4. Removal of the sol-
vent gave a crude yellowish mass. It was the purified over silica gel (70-
200 mesh) using 80% ethylacetate in petroleum ether to afford 7eas sticky
solid (0.52 g, 78%,). 1H NMR (CDCl3, 300 MHz) δ ppm: 1.76 (s, 3H,
Hz, H7). 13C NMR (CDCl3, 75 MHz) δ ppm: 18.22 (C13), 23.24 (C4 ),
0
0
0
25.83 (C5 ), 40.33 (C10 or 11), 40.35 (C11 or 10), 64.88 (C1 ), 114.95 (C3,5),
0
0
117.86 (C2 ), 119.25 (C8), 127.23 (C1), 129.41 (C2,6), 138.63 (C3 ),
140.93 (C7), 160.33 (C4), 167.57 (C9), 171.61 (C12). HRMS calcd for
C18H24N2O3 (M þ Na)þ, 339.1685; found, 339.1696. HPLC purity: 97%.
(E)-N-(2-Acetamidoethyl)-3-(4-methoxyphenyl)prop-2-enamide
(8a). Compound 8a was prepared from acid 6a according to pro-
cedure for 8e synthesis. White solid (0.046 g, 62%); mp 190-192 °C. IR
(KBr) ν cm-1: 3290 (N-H), 3084 (CdC-H), 1652 (CdO), 1619
0
0
H4 ), 1.81 (s, 3H, H5 ), 1.99 (s, 3H, H13), 3.16 (t, 2H, J = 6.4 Hz, H10),
3.50 (td, 2H, J = 6.4 Hz, HC11), 4.51 (d, 2H, J = 6.6 Hz, H1 ), 5.42 (th, 1H,
J = 6.6 Hz, J = 1.5 Hz, H2 ), 5.99 (s, 1H, NH), 6.54 (d, 1H, J = 15.9 Hz,
0
1
0
(CdC), 1559 (CdC arom.), 1227 (O-C arom), 975 (CHdCH). H
H8), 6.85 (d, 2H, J = 8.7 Hz, H3,5), 7.43 (d, 2H, J = 8.7 Hz, H2,6), 7.53 (d,
NMR (CDCl3, 300 MHz) δ ppm: 2.02 (s, 3H, H13), 3.50 (m, 4H, H10,11),
3.85 (s, 3H, MeO-), 6.31 (d, 1H, J = 15.6 Hz, H8), 6.50 (m, 2H, NH), 6.90
(d, 2H, J = 8.7 Hz, H3,5), 7.46 (d, 2H, J = 8.7 Hz, H2,6), 7.59 (d, 1H, J = 15.6
Hz, H7). 13C NMR (CDCl3, 75 MHz) δ ppm: 23.2 (C13), 40.3 (C10), 40.4
(C11), 55.36 (CH3), 114.2 (C3,5), 117.9 (C8), 127.3 (C1), 129.4 (C2,6),
140.9 (C7), 161.0 (C4), 167.5 (C9), 171.6 (C12). HRMS calcd for
C14H19N2O3 (MHþ), 263.1396; found, 263.1422. HPLC purity: 95%.
(E)-N-(2-Acetamidoethyl)-3-{4-[(E)-3,7-dimethylocta-2,6-dienyloxy]-
phenyl}prop-2-enamide (8f). Compound 8f was prepared from acid
6f according to the procedure for 8e synthesis. White solid (0.089 g,
83%); mp 155-157 °C. IR (KBr) ν cm-1: 3287 (N-H), 3087(CdC-
H), 2924 (C-H), 2856 (C-H), 1651 (CdO), 1623 (CdC), 1562
(CdC arom.), 1229 (O-C arom.), 974 (CHdCH). 1H NMR (CDCl3,
1H, J = 15.9 Hz, H7). 13C NMR (CDCl3, 300 MHz) δ ppm: 18.23 (s, 1C,
0
0
0
C4 ), 23.22 (C13), 25.82 (C5 ), 28.67 (C10), 39.92 (C11), 64.85 (C1 ),
0
115.17 (C3,5), 119.07 (C2 ), 122.10 (C8), 126.35 (C1), 130.27 (C2,6),
141.15 (C7), 161.24 (C4), 170.46 (C12), 190.00 (C9). MS (DCI, NH3,
pos.) m/z: 334.3 (MHþ), 351.3 (MNH4þ). HPLC purity: 100%.
(E)-S-2-Acetamidoethyl 3-(4-Methoxyphenyl)prop-2-enethioate
(7a). Compound 7a was prepared from acid 6a according to procedure
used for 7e synthesis. White solid (0.43 g, 77%); mp 96-97 °C. IR (KBr)
ν cm-1: 3268 (NH), 3168 (CH), 1640 (CO), 1548 (CdC arom.), 979
(HCdCH). 1H NMR (CDCl3, 300 MHz) δ ppm: 1.91 (s, 3H, C13), 3.15
(t, 2H, J = 6.4 Hz, CH2S), 3.50 (q, 2H, J = 5.9 Hz, H10), 3.85 (s, 3H, C13),
6.61 (d, 1H, J = 15.7 Hz, H8), 6.91 (d, 2H, J = 8.7 Hz, H3,5), 7.50 (d, 2H,
J = 8.7 Hz, H2,6), 7.59 (d, 1H, J = 15.7 Hz, H7). 13C NMR (CDCl3, 75
0
0
0
300 MHz) δ ppm: 1.36 (s, 3H, H4 ), 1.43 (s, 3H, H9 ), 1.48 (s, 3H, H10 ),
0
0
0
MHz) δ ppm: 23.4 (C13), 28.5 (C10), 40.0 (C11), 55.5 (C1 ), 114.5 (C3,
1.72 (s, 3H, H13), 1.85 (m, 4H, H5 ,6 ), 3.20 (t, 2H, J = 5.1 Hz, H10), 3.25
0
5), 122.2 (C7), 126.5 (C1), 130.24 (C2, 6), 141.20 (C8), 160.0 (C4), 165.6
(t, 2H, J = 5.1 Hz, H11), 4.28 (d, 2H, J = 6.5 Hz, H1 ), 4.83 (t, 1H, J = 6.2
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0
0
0
0
Hz, H7 ), 5.22 (t, 1H, J = 6.2 Hz, H2 ), 6.14 (d, 1H, J = 15.6 Hz, H8), 6.59
(d, 2H, J = 8.5 Hz, H3,5), 6.98 (t, 2H, J = 4.9 Hz, NH), 7.08 (t, 2H, J = 4.8
127.11 (C1), 129.66 (C2,6), 131.87 (C8 ), 138.58 (C12), 141.65 (C3 ),
142.70 (C7), 147.60 (C14), 152.89 (C10), 161.64 (C4), 164.87 (C9).
HRMS calcd for C24H28N2O2 (MHþ), 377.2253; found, 377.2229.
HPLC purity: 95%.
Hz, NH), 7.15 (d, 2H, J = 8.5 Hz, H2,6), 7.31 (d, 1H, J = 15.6 Hz, H7).
C NMR (CDCl3, 75 MHz) δ ppm: 16.67 (C4 ), 17.70 (C9 ), 23.16
13
0
0
0
0
0
(C10 ), 25.68 (C13), 26.26 (C6 ), 39.52 (C5 ), 40.01 (C10,11), 64.95
(E)-N-(3-Oxo-1,2-oxazolidin-4-yl)-3-(4-methoxyphenyl)prop-2-en-
amide (10a). A clean dry round-bottom flask (10 mL) was charged
with p-methoxycinnamic acid (0.06 g, 0.34 mmol, 1 equiv) and oxalyl
chloride (3 mL) at 0 °C and sealedwith anhydrous calcium chloride guard
tube. The reaction mixture was stirred at room temperature for 2 h, and
then, excess oxalyl chloride was removed in vacuo to give corresponding
acyl chloride quantitatively.
0
0
0
(C1 ), 114.96 (C3,5), 118.18 (C8), 119.05 (C2 ), 123.72 (C7 ), 127.24
0
0
(C1), 129.36 (C2,6), 131.82 (C8 ), 140.62 (C7), 141.58 (C3 ), 160.29
(C4), 167.65 (C9), 171.67 (C12). HRMS calcd for C23H32N2O3 (M þ
Na)þ, 407.2311; found, 407.2283. HPLC purity: 97%.
(E)-N-(Pyridin-2-yl)-3-[4-(3-methylbut-2-enyloxy)phenyl]prop-2-
enamide (9e). A clean dry round-bottom flask (25 mL) with a magnetic
stir bar was charged with carboxylic acid 6e (0.46 g, 2 mmol, 1 equiv),
2-aminopyridine (0.23 g, 2.4 mmol, 1.2 equiv), DMAP (0.26 g, 2.2 mmol,
In another round-bottom flask (25 mL) D-cycloserine (0.035 g, 0.34
mmol, 1 equiv) in dry dichloromethane (5 mL), N-methylpyrrolidine
(NMP) (1 mL, 9.6 mmol, 30 equiv) and bis-(trimethylsilyl)acetamide
(BSA) (0.17 g, 0.85 mmol, 2.5 equiv) were stirred under N2 at room
temperature for 1 h. The reaction mixture was cooled to 0 °C. Then,
pyridine (0.07 mL g, 6.8 mmol, 20 equiv) and acyl chloride dissolved in
dichloromethane (2 mL) were added to the reaction flask. The reaction
mixture was then stirred at room temperature for 12 h. Volatiles were
removed under reduced pressure. The residue dissolved in ethylacetate
(15 mL) and washed with dilute aqueous HCl (pH ca. 2, 3 ꢀ 5 mL) and
brine. The organic layer was dried over anhydrous Na2SO4 and then
evaporated in vacuo. The resulting brown solid was triturated with 10%
ethylacetate in hexanes, filtered, and dried to afford compound 10a as a
solid (0.063 g, 71%); mp 198-200 °C. [R]D þ 4.3; C = 0.01 g/cm3. IR
(KBr) ν cm-1: 3254 (N-H), 3058 (C-H), 1708 (CdO), 1651 (CdO),
1623 (CdC), 1604 (CdC arom.), 1512 (CdC arom.), 1256 (C-O
ether), 972 (CHdCH), 826 (C arom. 1-4). 1H NMR (MeOD-d4, 300
MHz) δppm: 3.80 (s, 3H, CH3), 4.11 (dd, 1H, J = 9.9 Hz, J= 8.6 Hz, H11),
4.66 (t, 1H, J = 8.5 Hz, H10), 5.03 (dd, 1H, J = 9.9 Hz,J0= 8.6 Hz, H11), 6.50
(d, 1H, J = 15.7 Hz, H8), 6.92 (d, 2H, J = 8.8 Hz, H3,5), 7.50 (d, 2H, J = 8.6
Hz, H2,6), 7.53 (d, 1H, J = 15.7 Hz, H7). 13C NMR(MeOD-d4, 75 MHz) δ
1.1 equiv), and EDC HCl (0.42 g, 2.2 mmol, 1.1 equiv) under argon. Dry
3
dichloromethane (15 mL) was added to it and stirred at room tempera-
ture for 24 h. TLC was monitored in ethylacetate. Dichloromethane was
removed under reduced pressure. Ethylacetate (60 mL) was added to the
crude yellow mass, and the solution was thoroughly washed with water
(30 mL ꢀ 3). The organic layer was then dried over MgSO4. Removal of
the solvent gave a crude yellowish mass. It was the purified over silica gel
(70-200 mesh) using 80% ethylacetate in petroleum ether to afford 9e as
a white solid (0.26 g, 42%); mp 130-132 °C). IR (KBr) ν cm-1: 3431
(NH), 3190 (HCdCH), 3007 (C-H), 1693 (CdO), 1630 (CdN),
1605 (CdC arom.), 1582 (CdC arom.), 1516 (CdC arom.), 1260 (C-
O), 987 (CHdCH), 823 (C arom.). H NMR (CDCl3, 300 MHz) δ
ppm: 1.68 (s, 3H, H4 ), 1.73 (s, 3H, H5 ), 4.48 (d, 2H, J = 6.8 Hz, H1 ),
5.42 (t, 1H, J = 6.8 Hz, H2 ), 6.52 (d, 1H, J = 15.6 Hz, H8), 6.82 (m, 1H,
H11), 6.84 (d, 2H, J = 8.5 Hz, H3,5), 7.13 (t, 1H, J = 7.3 Hz, H13), 7.42 (d,
2H, J = 8.5 Hz, H2,6), 7.68 (d, 1H, J = 15.6 Hz, H7), 7.89 (t, 1H, J = 7.3 Hz,
H14), 8.14 (d, 1H, J = 2.0 Hz, H11), 9.80 (s, 1H, NH). 13C NMR (CDCl3,
75 MHz) δ ppm: 14.19 (C4 ), 25.82 (C5 ), 64.92 (C1 ), 114.65 (C11),
115.04 (C3,5), 117.91 (C2 ), 119.21 (C8), 119.58 (C13), 127.61 (C1),
1
0
0
0
0
0
0
0
0
0
0
129.73 (C2,6), 138.67 (C3 ), 138.79 (C12), 142.88 (C7), 147.23 (C14),
ppm: 51.93 (C10), 54.45 (C1 ), 72.92 (C11), 113.98 (C3,5), 116.86 (C8),
151.95 (C10), 160.69 (C4), 164.79 (C9). HRMS calcd for C19H21N2O2
127.22 (C1), 129.24 (C2,6), 141.33 (C7), 161.40 (C4), 167.89 (C9), 170.56
(C12). HRMS calcd for C13H15N2O4 (MHþ), 263.1066; found, 263.1072.
HPLC purity: 96%.
(MHþ), 309.1603; found, 309.1591. HPLC purity: 95%.
(E)-N-(Pyridin-2-yl))-3-(4-mehoxyphenyl)prop-2-enamide (9a). Com-
pound 9a was prepared from acid 6a according to procedure used for 9e
synthesis (0.37 g, 73%). White solid; mp 93-95 °C. IR (KBr) ν cm-1: 3432
(NH), 3194 (HCdCH), 3008 (C-H), 1693 (CdO), 1630 (CdN), 1605
(CdC arom.), 1582 (CdC arom.), 1516 (CdC arom.), 1260 (C-O), 988
(CHdCH), 824 (C arom.). 1H NMR (CDCl3, 300 MHz) δ ppm: 3.83 (s,
Representative Procedure: Preparation of (E)-N0-{3-[4-(3-Methyl-
but-2-enyloxy)phenyl]propenoyl}isonicotinohydrazide (11e). A clean
round-bottom flask (25 mL) was charged with 6e (0.2 g, 0.86 mmol,
1 equiv), HBTU (0.49 g, 1.29 mmol, 1.5 equiv), and INH (0.18 g, 1.29
mmol, 1.5 equiv) in dry DMF (10 mL). Diisopropylethylamine (0.3 mL,
1.72 mmol, 2 equiv) was added to it, and the reaction mixture was stirred
at room temperature for 24 h. Dimethylformamide was removed under
vacuum, and brine (20 mL) was added to the residue. The aqueous phase
was extracted with ethyl acetate (40 mL ꢀ 3). The combined organic
phases were dried over anhydrous Na2SO4. Removal of the solvent under
reduced pressure gave a yellow residue that was purified over silica gel
using 5% methanol in dichloromethane to afford compound 11e. White
solid (0.206 g, 68%); mp 176-178 °C. IR (KBr) ν cm-1: 3242 (N-H),
3019 (CdC-H), 1679 (CdO), 1634 (CdC), 1605 (CdC), 1587
(CdC arom.), 1495 (CdC arom.), 1176 (O-C), 1466 (CH2), 981
(CHdCH), 837 (C arom 1-4). 1H NMR (CDCl3, 300 MHz) δ ppm:
0
3H, H1 ), 6.50 (d, 1H, J = 15.5 Hz, H8), 6.88 (d, 2H, J = 8.7 Hz, H3,5), 7.10
(dd, 1H, J = 7.3 Hz, J = 5.0 Hz, H13), 7.45 (d, 2H, J = 8.7 Hz, H2,6), 7.75 (d,
1H, J = 15.5 Hz, H7), 7.75 (m, 1H, H12), 8.30 (d, 1H, J = 4.9 Hz, H14), 8.40
(d, 1H, J = 8.4 Hz, H11), 9.28 (s, 1H, NH). 13C NMR (CDCl3, 75 MHz) δ
0
ppm: 55.3 (C1 ), 114.3 (C11), 114.6 (C3,5), 118.1 (C8), 119.6 (C13), 127.2
(C1), 129.7 (C2,6), 138.5 (C12), 142.6 (C7), 147.6 (C14), 152.0 (C10), 161.3
(C4), 164.6 (C9). HRMS calcd for C15H15N2O2 (MHþ), 255.1134; found,
255.1141. HPLC purity: 95%.
(E)-N-(Pyridin-2-yl)-3-{4-[(E)-3,7-dimethylocta-2,6-dienyloxy]phenyl}-
prop-2-enamide (9f). Compound 9f was prepared from acid 6f according
to procedure used for 9e synthesis (0.72 g, 48%). White solid; mp 98-
100 °C. IR (KBr) ν cm-1: 3426 (NH), 3187 (HCdCH), 3008 (C-H),
1697 (CdO), 1634 (CdN), 1604 (CdC arom.), 1579 (CdC arom.),
1514 (CdC arom.), 1261 (C-O), 981 (CHdCH), 827(C arom.). 1H
0
0
0
0
1.77 (bs, 3H, H4 ), 1.82 (bs, 3H, H5 ), 4.55 (d, 2H, J = 6.6 Hz, H1 ), 5.49 (t,
0
1H, J = 6.6 Hz, J = 1.3Hz, H2 ), 6.51 (d, 1H, J = 15.6 Hz, H8), 6.88 (d, 2H,
J = 8.7 Hz, H3,5), 7.40 (d, 2H, J = 8.7 Hz, H2,6), 7.66 (d, 1H, J = 15.6 Hz,
H7), 7.71 (d, 2H, J = 5.6 Hz, H12,15), 8.73 (d, 2H, J = 5.6 Hz, H13,14), 9.59
(bs, 1H, NH), 10.34 (bs, 1H, NH). 13C NMR0(CDCl3, 75 MHz) δ ppm:
0
0
NMR (CDCl3, 300 MHz) δ ppm: 1.38 (s, 3H, H4 ), 1.45 (s, 3H, H9 ),
0
0
0
0
1.51 (s, 3H, H10 ), 1.87 (m, 4H, H5 ,6 ), 4.34 (d, 2H, J = 6.5 Hz, H1 ), 4.87
0
0
0
0
0
18.11 (C4 ), 25.62 (C5 ), 65.04 (C1 ), 114.12 ( C8), 115.17 (C3,5), 119.34
(t, 1H, J = 6.6 Hz, H7 ), 5.25 (t, 1H, J = 6.5 Hz, H2 ), 6.25 (d, 1H, J = 15.5
Hz, H8), 6.66 (d, 2H, J = 8.7 Hz, H3,5), 6.83 (dd, 1H, J = 7.2 Hz, J0 = 5.5
Hz, H13), 7.21 (d, 2H, J = 8.7 Hz, H2,6), 7.52 (d, 1H, J = 15.5 Hz, H7), 7.53
(m, 1H, H12), 8.10 (d, 1H, J = 4.5 Hz, H14), 8.20 (d, 1H, J = 8.4 Hz, H11),
0
0
(C12,15), 121.0 (C2 ), 126.88 (C1), 129.66 (C2,6), 138.33 (C3 ), 138.57
(C11), 143.27 (C7), 150.61 (C13,14), 160.96 (C4), 162.36 (C10), 163,93
(C9). HRMS calcd for C20H22N3O3 (MHþ), 352.1685; found, 352.1661.
HPLC purity: 95%.
9.22 (s, 1H, NH). 13C NMR (CDCl3, 75 MHz) δ ppm:16.71 (C4 ), 17.72
0
(C9 ), 25.70 (C10 ), 26.27 (C6 ), 39.53 (C5 ), 65.01 (C1 ), 114.77 (C11),
(E)-N0-[3-(4-Methoxyphenyl)propenoyl]isonicotinohydrazide (11a).
Compound 11a was prepared from acid 6a according to procedure used
0
0
0
0
0
0
0
115.07 (C3,5), 118.04 (C8), 119.07 (C2 ), 119.53 (C13), 123.73 (C7 ),
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for 11e synthesis. White solid (0.95 g, 74%); mp 242-245 °C. IR (KBr) ν
cm-1: 3208 (N-H), 3028 (CdC-H), 2837 (C-H), 1688 (CdO),
1630 (CdC), 1587 (CdC arom.), 1498 (CdC arom.), 1254 (O-C),
981 (CHdCH), 826 (C arom. 1-4). 1H NMR (DMSO-d6, 300 MHz) δ
J = 6.7 Hz, H7 ), 5.46 (t, 1H, J = 6.6 Hz, H2 ), 6.54 (d, 1H, J = 15.7 Hz, H8),
6.83 (d, 2H,J = 8.0 Hz, H3,5), 7.35 (d, 2H, J = 8.0 Hz, H2,6), 7.57 (d, 1H, J =
15.4 Hz, H7), 7.71 (d, 2H, J = 4.6 Hz, H12,15), 8.64 (d, 2H, J = 4.3 Hz, H13-
14), 10.17 (s, 1H, NH), 11.04 (m, 1H, NH). 13C NMR (CDCl3, 75 MHz)
0
0
0
0
0
0
ppm: 3.80 (s, 3H, H1 ), 6.62 (d, 1H,J =16.2 Hz, H8), 7.0 (d, 2H, J = 8.5 Hz,
δ ppm: 16.72 (C9 ), 17.73 (C10 ), 25.71 (C4 ), 26.30 (C6 ), 39,55 (C5 ),
0
0
0
H2,6), 7.55 (d, 1H, J = 16.2 Hz, H7), 7.58 (d, 2H, J = 8.5 Hz, H3,5), 7.81 (d,
65.0 (C1 ), 114.36 (C8), 115.02 (C3,5), 118.7 (C7 ), 121.37 (C2 ), 123.73
0
0
2H, J = 4.2 Hz, H12,15), 8.70 (d, 2H, J = 4.2 Hz, H13,14), 10.24 (s, 1H, NH),
(C12,15), 126.86 (C1), 129.72 (C2,6), 131.86 (C8 ), 138.67 (C3 ), 141.72
(C11), 143.03 (C7), 150.30 (C13,14), 160.79 (C4), 163.41 (C10), 165.19
(C9). HRMS calcd for C25H30N3O3 (MHþ), 420.2296; found, 420.2287.
HPLC purity: 95%.
10.83 (s, 1H, NH). 13C NMR (DMSO-d6, 75 MHz) δ ppm: 55.77 (C1 ),
0
114.93 (C3,5), 117.09 (C8), 121.81 (C12,15), 127.54 (C1), 129.90 (C2,6),
139.93 (C11), 140.74 (C7), 150.89 (C13,14), 161.16 (C4), 164.38 (C10),
165.10 (C9). HRMS calcd for C16H15N3O3 (MHþ), 298.1192; found,
298.1212. HPLC purity: 95%.
(2E,N0,E)-3-[4-(3-Methylbut-2-enyloxy)phenyl]-N0-(phthalazin-1-(2H)-
ylidiene)acrylo Hydrazide (12e). A clean dry round-bottom flask (10 mL)
with a magnetic stir bar was charged with carboxylic acid (6e, 0.64 g, 1.7
mmol, 1 mmol), 1-hydrazinophthalazine hydrochloride (0.5 g, 2.53 mmol,
(E)-N0-[3-(4-Trifluoromethoxyphenyl)propenoyl]isonicotinohydra-
zide (11b). Compound 11b was prepared from acid 6b according to
procedure used for 11e synthesis. White solid (0.24 g, 81%); mp 196-
198 °C. IR (KBr) ν cm-1: 3197 (NH), 3018 (CdCH), 1638 (CdO),
1587 (CdC arom.), 1554 (CdC arom.), 1271 (C-O), 1218 (C-F),
971 (CHdCH), 843 (C arom. 1-4). 1H NMR (DMSO-d6, 300 MHz)
δ ppm: 6.79 (d, 1H, J = 15.9 Hz, H8), 7.43 (d, 2H, J = 8.3 Hz, H3,5), 7.63
(d, 1H, J = 15.9 Hz, H7), 7.78 (d, 2H, J = 8.7 Hz, H2,6), 7.82 (d, 2H, J =
5.9 Hz, H12,15), 8.78 (d, 2H, J = 5.8 Hz, H13,14), 10.43 (m, 1H, NH),
10.83 (m, 1H, NH). 13C NMR (DMSO-d6, 75 MHz) δ ppm: 120.46 (q,
J = 255.0 Hz, CF3), 120.90 (C8), 121.81 (C12,15), 121.86 (C3,5), 130.17
(C2,6), 134.29 (C1), 139.40 (C7), 139.84 (C11), 149.54 (C4), 150.88
(C13,14), 164.35 (C10), 164.48 (C9). 19F NMR (DMSO-d6, 282 MHz) δ
ppm: -56.76 (s, 3F, CF3O-). HRMS calcd for C16H13F3N3O3
(MHþ), 352.0928; found, 352.9022. HPLC purity: 98%.
1.5 equiv), HOBt (0.27 g, 2 mmol, 1.2 equiv), and EDC HCl (0.48 g, 2.53
3
mmol, 1.5 equiv) under argon. Dry dichloromethane (15 mL) was added to
it and stirred. Triethylamine (0.47 mL, 3.4 mmol, 2 equiv) was added to
the reaction mixture andstirredatroom temperature for 24 h, thenpoured
into aqueous saturated NH4Cl solution, extracted with dichloromethane
(60 mL ꢀ 3), and was thoroughly washed with water (30 mL ꢀ 3). The
organic layer was then dried over MgSO4. Removal of the solvent gave a
crude yellowish mass thatwas filtered off to afford 12e. Yellow solid (0.5 g,
77%); mp 175-177 °C. IR (KBr) ν cm-1: 3205 (NH), 2980 (CdCH),
2921 (C-H), 1644 (CdO), 1604 (CdC), 1572 (CdC arom.), 1547
(CdC arom.), 1222 (C-O), 984 (CHdCH), 827 (C arom. 1-4). 1H
NMR (DMSO-d6, þ few drops of CF3COOD, 300 MHz) δ ppm: 1.70 (s,
0
0
0
3H, H4 ), 1.72 (s, 3H, H5 ), 4.56 (d, 2H, J = 6.7 Hz, H1 ), 5.42 (t, 1H, J =
6.6 Hz, H2 ), 6.68 (d, 1H, J = 15.9 Hz, H8), 670 (d, 2H, J = 8.7 Hz, H3,5),
(E)-N0-[3-(4-Ethoxyphenyl)propenoyl]isonicotinohydrazide (11c).
Compound 11c was prepared from acid 6c according to procedure
used for 11e synthesis. White solid (0.19 g, 65%); mp 215-217 °C. IR
(KBr) ν cm-1: 3277 (NH), 3028 (CdCH), 1662 (CdO), 1628
0
7.60 (d, 2H, J = 8.7 Hz, H2,6), 7.62 (d, 1H, J = 15.8 Hz, H7), 8.19 (td, 1H,
J = 7.1 Hz, J0 = 1.7Hz, H13), 822 (dd, 1H, J = 7.6 Hz, J0 = 1.8Hz, H12), 8.28
(td, 1H, J = 7.4 Hz, J0 = 1.7 Hz, H14), 8.73 (d, 1H, J = 7.6 Hz, H15), 9.14 (s,
1H, H17). 13C NMR (DMSO-d6, þ few drops of CF3COOD, 75 MHz) δ
(CdCH), 1604 (CdC arom.), 1513 (CdC arom.), 1260 (C-O).
1
0
0
0
0
H NMR (DMSO-d6, 300 MHz) δ ppm: 1.40 (t, 3H, J = 6.9 Hz, H2 ),
ppm: 18.14 (C4 ), 25.57 (C5 ), 64.89 (C1 ), 115.53 (C3,5), 116.40 (C8),
0
0
4.13 (q, 2H, J = 6.9 Hz, H1 ), 6.66 (d, 1H, J = 15.8 Hz, H8), 7.04 (d, 2H,
117.33 (C11), 119.93 (C2 ), 124.66 (C15), 127.14 (C1), 128.13 (C16),
0
J = 8.7 Hz, H3,5), 7.59 (d, 1H, J = 15.4 Hz, H7), 7.63 (d, 2H, J = 8.6 Hz,
H2,6), 7.87 (d, 2H, J = 6.1 Hz, H12,15), 8.84 (d, 2H, J = 6.0 Hz, H13,14),
10.29 (s, 1H, NH), 10.89 (s, 1H, NH). 13C NMR (DMSO-d6, 75 MHz)
129.18 (C12), 129.98 (C2,6), 134.96 (C13), 136.78 (C14), 137.88 (C3 ),
141.72 (C7), 145.88 (C17), 152.61 (C4), 160.71 (C10), 166.04 (C9).
HRMS calcd for C22H23N4O2 (MHþ), 375.1836; found, 375.1821.
HPLC purity: 95%.
0
0
δ ppm: 14.11 (C2 ), 62.78 (C1 ), 114.40 (C3,5), 116.04 (C8), 120.87
(C12,15), 126.44 (C1), 128.96 (C2,6), 138.98 (C11), 139.81 (C7), 149.96
(C13,14), 159.50 (C4), 163.42 (C10), 164.15 (C9). HRMS calcd for
C17H18N3O3 (MHþ), 312.1353; found, 312.1348. HPLC purity: 95%.
(E)-N0-[3-(4-(20,20,20-Trifluoroethoxyphenyl]propenoyl]isonicotinohydr-
azide (11d). Compound 11d was prepared from acid 6d according to proce-
dure used for 11e synthesis. White solid (0.23 g, 74%); mp 208-210 °C.
IR (KBr) ν cm-1: 3225 (NH), 3027 (CdCH), 2854 (C-H), 1635
(2E,N0,E)-3-(4-Methoxyphenyl)-N0-[phthalazin-1-(2H)-ylidiene]acrylo-
hydrazide (12a). Compound 12a was prepared from acid 6a according to
procedure used for 12e synthesis. Yellow solid (0.42 g, 78%); mp 194-
196 °C. IR (KBr) ν cm-1: 3199 (NH), 3072 (CdCH), 2996 (C-H),
1643 (CdO), 1604 (CdC arom.), 1546 (CdC arom.), 1259 (C-O), 982
(CHdCH), 828 (C arom. 1-4). 1H NMR (DMSO-d6, þ few drops of
0
CF3COOD, 300 MHz) δ ppm: 3.72 (s, 3H, C1 ), 6.61 (d, 2H, J = 15.9 Hz,
(CdO), 1604 (CdC arom.) (CdC arom.), 1243 (C-O), 1172 (C-F).
H NMR (DMSO-d6, 300 MHz) δ ppm: 4.83 (q, 2H, J = 8.9 Hz, H1 ),
6.66 (d, 1H, J = 15.8 Hz, H8), 7.13 (d, 2H, J = 8.8 Hz, H3,5), 7.56 (d, 1H,
J = 15.8 Hz, H7), 7.63 (d, 2H, J = 8.8 Hz, H2,6), 7.81 (d, 2H, J = 6.1 Hz,
H12,15), 8.78 (d, 2H, J = 6.0 Hz, H13,14). 13C NMR (DMSO-d6, 75 MHz)
H8), 6.93 (d, 2H, J = 8.7 Hz, H2,6), 7.55 (d, 2H, J = 9.0 Hz, H3,5), 7.56 (d,
1H, J= 15.1 Hz, H7), 8.13 (td, 1H, J=7.8Hz, J0 =1.7Hz, H13), 8.18 (td, 1H,
J = 7.5 Hz, J0 = 2.2 Hz, H14), 8.22 (dd, 1H, J = 7.4 Hz, J0 = 2.2 Hz, H12), 8.62
(dd, 1H, J=7.7Hz, J0 =1.7Hz, H15), 9.08(s, 1H, H17). 13C NMR (DMSO-
1
0
0
d6, þ few drops of CF3COOD, 75 MHz) δ ppm: 55.61 (C1 ), 114.96
0
δ ppm: 65.04 (q, J = 34.5 Hz, C1 ), 115.81 (C3,5), 118.12 (C8), 118.89
(C3,5), 116.54 (C8), 118.63 (C11), 124.61 (C15), 127.29 (C1), 128.14
(C16), 129.26 (C12), 130.07 (C2,6), 135.06 (C13), 136.86 (C14), 141.70
(C7), 145.93 (C17), 152.65 (C4), 161.47 (C10), 166.07 (C9). HRMS calcd
for C18H16N4O2 (MHþ), 321.1351; found, 321.1370. HPLC purity: 95%.
(2E,N0,E)-3-(4-Trifluoromethoxyphenyl)-N0-[phthalazin-1-(2H)-yli-
diene]acrylohydrazide (12b). Compound 12b was prepared from acid
6b according to procedure used for 12e synthesis. Yellow solid (0.41 g,
65%); mp 210-212 °C. IR (KBr) ν cm-1: 3204 (NH), 3070 (CdCH),
1658 (CdO), 1572 (CdC arom.), 1548 (CdC arom.), 1266 (C-O),
1218 (C-F), 986 (CHdCH), 841 (C arom. 1-4). 1H NMR (DMSO-
d6, þ few drops of CF3COOD, 300 MHz) δ ppm: 6.87 (d, 1H, J = 15.9
Hz, H8), 7.42 (d, 2H, J = 8.3 Hz, H3,5), 7.72 (d, 1H, J = 15.9 Hz, H7), 7.82
(d, 2H, J = 8.7 Hz, H2,6), 8.21 (td, 1H, J = 7.1 Hz, J0= 1.4 Hz, H13), 8.25
(d, 1H, J = 7.2 Hz, H12), 8.30 (td, 1H, J = 7.3 Hz, J0= 1.4 Hz, H14), 8.71
0
(C12,15), 124.41 (q, 1C, J = 276 Hz, C2 ), 129.09 (C1), 129.93 (C2,6),
139.91 (C11), 140.30 (C7), 150.89 (C13,14), 158.60 (C4), 164.36 (C10),
164,93 (C9). 19F NMR (DMSO-d6, 282 MHz) δ ppm: -72,48 (t, 3F, J =
8.8 Hz, CF3-). HRMS calcd for C17H15F3N3O3 (MHþ), 366.1050;
found, 366.1066. HPLC purity: 95%.
(E)-N0-[3-{4-[(E)-3,7-Dimethylocta-2,6-dienyloxy]phenyl}propenoyl]-
isonicotinohydrazide (11f). Compound 11f was prepared from acid 6f
according to procedure used for 11e synthesis. White solid (0.22 g, 61%);
mp 110-112 °C. IR (KBr) ν cm-1: 3249 (NH), 2969 (CdCH), 2923
(C-H), 1678 (CdCH), 1646 (CdO), 1604 (CdC arom.), 1513 (CdC
1
arom.), 1236 (C-O), 997 (CHdCH), 844 (C arom. 1-4). H NMR
0
0
(CDCl3, 300 MHz) δ ppm: 1.60 (s, 3H, H9 ), 1.67 (s, 3H, H10 ), 1.73 (s,
0
0
0
0
3H, H4 ), 2.10 (m, 4H, H5 ,6 ), 4.53 (d, 2H, J = 6.6 Hz, H1 ), 5.07 (t, 1H,
1458
dx.doi.org/10.1021/jm101510d |J. Med. Chem. 2011, 54, 1449–1461

Journal of Medicinal Chemistry
ARTICLE
(d, 1H, J = 7.7 Hz, H15), 9.71 (s, 1H, H17). 13C NMR (DMSO-d6, þ few
drops of CF3COOD, 75 MHz) δ ppm: 118.10 (C11), 119.91 (C8),
119.95 (q, 1C, J = 255 Hz, CF3), 121.27 (C3,5), 124.11 (C15), 127.11
(C1), 128.75 (C12), 129.10 (C2,6), 133.47 (C16), 134.56 (C13), 136.31
(C14), 139.65 (C7), 145.44 (C17), 149.35 (C4), 152.11 (C10), 164.96
(C9). 19F NMR (DMSO-d6, þ few drops of CF3COOD, 282 MHz) δ
ppm: -57.71 (s, 3F). HRMS calcd for C18H14F3N4O2 (MHþ), 375.1059;
found, 375.1069. HPLC purity: 95%.
(E)-3-[4-(3-Methylbut-2-enyloxy)styryl]-[1,2,4]triazolo[3,4-R]phth-
alazine (13e). A clean dry round-bottom flask (25 mL) with a magnetic
stir bar was charged with carboxylic acid (6e, 0.46 g, 1.3 mmol, 1 equiv),
1-hydrazinophthalazine hydrochloride (0.25 g, 1.3 mmol, 1 equiv),
HOBt (0.25 g, 1,3 mmol, 1 equiv), and EDC HCl (0.25 g, 1.3 mmol,
3
1 equiv) under argon. Dry acetonotrile (10 mL) was added to it and
stirred. Triethylamine (0.24 mL, 1.72 mmol, 1.1 equiv) was added to the
reaction mixture and refluxed for 48 h. TLC was monitored in ethyl-
acetate. Acetonitrile was removed under reduced pressure. Ethylacetate
(40 mL) was added to the crude yellow mass, and the solution was
thoroughly washed with water (30 mL ꢀ 3). The organic layer was then
dried over MgSO4. Removal of the solvent gave a crude yellowish mass.
It was then purified over silica gel (70-200 mesh) using 80% ethyl-
acetate in petroleum ether to afford 13e. Yellow solid (0.25 g, 52%); mp
170-172 °C. IR (KBr) ν cm-1: 2964 (CdC-H), 1638 (CdC), 1603
(CdC arom.), 1515 (CdC arom.), 1246 (O-C), 966 (CHdCH
(2E,N0,E)-3-(4-Ethoxyphenyl)-N0-[phthalazin-1-(2H)-ylidiene]acryl-
ohydrazide (12c). Compound 12c was prepared from acid 6c accord-
ing to procedure used for 12e synthesis. Yellow solid (0.32 g, 55%); mp
225-227 °C. IR (KBr) ν cm-1: 3204 (NH), 3073 (CdCH), 2979 (C-
H), 1643 (CdO), 1604 (CdC arom.), 1571 (CdC arom.), 1510
1
(CdC arom.), 1259 (C-O). H NMR (DMSO-d6, þ few drops of
0
CF3COOD, 300 MHz) δ ppm: 1.24 (t, 3H, J = 6.9 Hz, H2 ), 3.98 (q, 2H,
0
J = 6.9 Hz, H1 ), 6.58 (d, 1H, J = 15.9 Hz, H8), 6.90 (d, 2H, J = 8.6 Hz,
1
H3,5), 7.52 (d, 2H, J = 8.7 Hz, H2,6), 7.55 (d, 1H, J = 15.7 Hz, H7), 8.11
(td, 1H, J = 7.1 Hz, J0= 1.6 Hz, H13), 8.14 (dd, 1H, J = 7.5 Hz, J0= 1.7 Hz,
H12), 8.20 (td, 1H, J = 7.3 Hz, J0= 1.6 Hz, H14), 8.61 (d, 1H, J = 7.6 Hz,
H15), 9.05(s, 1H, H17). 13CNMR(DMSO-d6, þ few drops of CF3COOD,
trans), 834 (C arom. 1-4). H NMR (CDCl3 300 MHz) δ ppm:
0
0
0
1.77 (bs, 3H, H4 ), 1.81 (bs, 3H, H5 ), 4.56 (d, 2H, J = 6.7 Hz, H1 ), 5.51
0
(t, 1H, J = 6.8 Hz, H2 ), 6.96 (d, 2H, J = 8.8 Hz, H3,5), 7.37 (d, 1H, J =
16.5 Hz, H8), 7.60 (d, 2H, J = 8.7 Hz, H2,6), 7.81 (td, 1H, J = 6.8 Hz, J0 =
1.0 Hz, H13), 7.94 (d, 1H, J = 7.6 Hz, H12), 7.96 (td, 1H, J = 7.3 Hz, J0 =
1.2 Hz, H14), 8.11 (d, 1H, J = 16.5 Hz, H7), 8.67 (s, 1H, H17), 8.70 (d,
0
0
75 MHz) δ ppm: 14.29 (C2 ), 63.15 (C1 ), 114.82 (C3,5), 115.79 (C8),
118.07 (C11), 124.02 (C15), 126.54 (C1), 127.61 (C16), 128.75 (C12),
129.55 (C2,6), 134.55 (C13), 136.36 (C14), 141.22 (C7), 145.44 (C17),
152.11 (C4), 160.24 (C10), 165.49 (C9). HRMS calcd for C19H19N4O2
(M þ H)þ, 335.1512; found, 335.1508. HPLC purity: 95%.
1H, J = 7.8 Hz, H15). 13C NMR (CDCl3 75 MHz) δ ppm: 18.25 (C4 ),
0
0
0
0
25.85 (C5 ), 64.90 (C1 ), 108.03 (C8), 115.04 (C3,5), 119.40 (C2 ),
123.14 (C11), 123.42 (C15), 123.54 (C16), 128.08 (C12), 128.68 (C1),
(2E,N0,E)-3-[4-(20,20,20-Ttrifluoroethoxy)phenyl]-N0-[phthalazin-1-
(2H)-ylidiene]acrylo Hydrazide (12d). Compound 12d was prepared
from acid 6d according to procedure used for 12e synthesis. Yellow solid
(0.43 g, 66%); mp 254-256 °C. IR(KBr) ν cm-1: 3152 (NH), 3049 (C-
H), 1650 (CdO), 1605 (CdC arom.), 1550 (CdC arom.), 1246 (C-
O), 1174 (C-F). 1H NMR (DMSO-d6, þ few drops of CF3COOD, 300
MHz) δ ppm: 4.80 (q, 2H, J = 8.8 Hz, CH2), 6.71 (d, 1H, J = 15.9 Hz, H8),
7.12 (d, 2H, J = 8.7 Hz, H3,5), 7.61 (d, 1H, J = 15.2 Hz, H7), 7.65 (d, 2H, J =
8.3 Hz, H2,6), 8.20 (td, 1H, J = 7.0 Hz, J0= 1.4 Hz, H13), 8.23 (dd, 1H, J =
6.9 Hz, J0= 1.8 Hz, H12), 8.29 (td, 1H, J = 7.3 Hz, J0= 1.7 Hz, H14), 8.72 (d,
1H, J =7.7 Hz, H15), 9.15(s, 1H, H17). 13C NMR (DMSO-d6, þ few drops
of CF3COOD, 75 MHz) δ ppm: 65.05 (q, J = 30.6 Hz, CH2), 115.84
(C3,5), 117.59 (C8), 118.63 (C11), 124.33 (q, J = 278.0 Hz, CF3), 124.69
(C15), 128.13 (C1), 128.81 (C16), 129.29 (C12), 130.01(C2,6), 135.10
(C13), 136.89 (C14), 141.10 (C7), 145.90 (C17), 152.59 (C4), 158.83
(C10), 165.81 (C9). 19F NMR (DMSO-d6, þ few drops of CF3COOD,
282 MHz) δ ppm: -72,68 (t, 3F, J = 8,8 Hz). HRMS calcd for
C19H16F3N4O2 (MHþ), 389.1228; found, 389.1225. HPLC purity: 95%.
(2E,N0,E)-3-{4-(E)-[3,7-Dimethylocta-2,6-dienyloxy]phenyl}-N0-
[phthalazin-1-(2H)-ylidiene]acrylo Hydrazide (12f). Compound
12f was prepared from acid 6f according to procedure used for 12e
0
128.82 (C2,6), 130.90 (C13), 134.18 (C14), 136.17 (C7), 138.58 (C3 ),
142.71 (C9), 147.54 (C17), 149.04 (C10), 159.89 (C4). HRMS calcd for
C22H20N4O (MHþ), 357.1715; found, 357.1748. HPLC purity: 95%.
(E)-3-(4-Methoxystyryl)-[1,2,4]triazolo[3,4-R]phthalazine (13a). Com-
pound 13a was prepared from acid 6a according to procedure used for 13e
synthesis. Yellow solid (0.24 g, 58%); mp 193-195 °C. IR (KBr) ν cm-1
:
3028 (CdC-H), 2837 (C-H), 1642 (CdC), 1604 (CdC arom.), 1517
(CdC arom.), 1255 (O-C arom), 1246 (O-C), 983 (CHdCH), 821 (C
1
0
arom. 1-4). HNMR(CDCl3, 300MHz) δppm: 3.84 (s, 3H, H1 ),6.92(d,
2H, J = 8.7 Hz, H3,5), 7.33 (d, 1H, J = 16.6 Hz, H8), 7.58 (d, 2H, J = 8.8 Hz,
H2,6), 7.79 (td, 1H, J=7.4Hz, J0 =0.6Hz, H13), 7.82 (d, 1H, J=7.6Hz, H12),
7.95 (td, 1H, J = 7.0 Hz, J0 = 0.7 Hz, H14), 8.06 (d, 1H, J = 16.6 Hz, H7), 8.65
(d, 1H, J = 7.4 Hz, H15), 8.65 (s, 1H, H17). 13C NMR (CDCl3 75 MHz) δ
0
ppm: 55.42 (C1 ), 108.22 (C8), 114.25 (C3,5), 123.05 (C11), 123.28 (C15),
123.51 (C16), 128.11 (C12), 128.72 (C1), 128.80 (C2,6), 130.85 (C13),
134.13 (C14), 135.73 (C7), 142.75 (C9), 147.50 (C17), 148.90 (C10), 160.40
(C4). HRMS calcd for C18H15N4O (MHþ), 303.1256; found, 303.1246.
HPLC purity: 97%.
(E)-3-(4-Trifluoromethoxystyryl)-[1,2,4]triazolo[3,4-R]phthalazine
(13b). Compound 13b was prepared from acid 6b according to
procedure used for 13e synthesis. Yellow solid (0.40 g, 87%); mp 190-
192 °C. IR (KBr) ν cm-1: 3043 (C-H), 1628 (CdC), 1603 (CdC
arom.), 1512 (CdC arom.), 1301 (C-F), 1219 (C-O), 988 (CHdCH),
839 (C arom. 1-4). 1H NMR (DMSO-d6, 300 MHz) δ ppm: 7.34 (d, 2H,
J = 8.0 Hz, H3,5), 7.53 (d, 1H, J = 16.6 Hz, H8), 7.88 (d, 2H, J = 8.7 Hz,
H2,6), 7.91 (td, 1H, J = 7.4 Hz, J0 = 1.2 Hz, H13), 8.0 (d, 1H, J = 16.6 Hz,
H7), 8.04 (td, 1H, J = 7.4 Hz, J0 = 1.2 Hz, H14), 8.21 (d, 1H, J = 7.6 Hz,
H12), 8.51 (d, 1H, J = 7.4 Hz, H15), 9.11 (s, 1H, H17). 13C NMR (DMSO-
d6, 75 MHz) δ ppm: 112.85 (C8), 120.57 (q, J = 255.7 Hz, CF3), 121.70
(C3,5), 122.61 (C15), 123.22 (C11), 123.53 (C16), 129.52 (C12), 129.55
(C2,6), 131.51 (C13), 133.48 (C7), 134.49 (C14), 135.53 (C1), 143.01
(C9), 148.01 (C10), 148.96 (C17), 149.05 (C4). 19F NMR (DMSO-d6, 282
MHz) δ ppm: -56,67 (s, 3F, F1). HRMS calcd for C18H12F3N4O
(MHþ), 357.0951; found, 357.0963. HPLC purity: 97%.
synthesis. Yellow solid (0.38 g, 51%); mp 164-166 °C. IR (KBr) ν cm-1
:
3204 (NH); 2968 (CdCH), 2920 (C-H), 1649 (CdO), 1604 (CdC),
1573 (CdC arom.), 1547 (CdC arom.), 1221 (C-O), 984 (CHdCH),
825 (C arom.). 1H NMR (DMSO-d6, þ few drops of CF3COOD, 300
0
0
0
MHz) δ ppm: 1.56 (s, 3H, H9 ), 1.62 (s, 3H, H10 ), 1.72 (s, 3H, H4 ), 2.05
0
0
0
0
(m, 4H, H5 ,6 ), 4.61 (d, 2H, J = 6.2 Hz, H1 ), 5.07 (t, 1H, J = 6.6 Hz, H7 ),
0
5.44 (t, 1H, J = 6.3 Hz, H2 ), 6.67 (d, 1H, J = 15.8 Hz, H8), 7,02 (d, 2H, J =
8.5 Hz, H3,5), 7.63 (d, 2H, J = 8.9 Hz, H2,6), 7.64 (d, 1H, J = 15.3 Hz, H7),
8.24 (t, 1H, J = 7.0 Hz, H13), 8.27 (d, 1H, J = 6.6 Hz, H12), 8.32 (t, 1H, J =
7.2 Hz, H14), 8.69 (d, 1H, J = 7.1 Hz, H15), 9.18 (s, 1H, H17). 13C
NMR (DMSO-d6, þ few drops of CF3COOD, 75 MHz) δ ppm: 16.75
0
0
0
0
0
0
(C9 ), 17.92 (C10 ), 25.85 (C4 ), 26.24 (C6 ), 39.32 (C5 ), 65.02 (C1 ),
0
115.67 (C3,5), 116.40 (C8), 118.65 (C11), 119.85 (C2 ), 124.18 (C15),
0
124.59 (C7 ), 127.16 (C1), 128.16 (C16), 129.34 (C12), 130.08 (C2,6),
(E)-3-(4-Ethoxystyryl)-[1,2,4]triazolo[3,4-R]phthalazine (13c). Com-
pound 13c was prepared from acid 6c according to procedure used for 13e
0
0
131.48 (C8 ), 135.15 (C13), 136.94 (C14), 141.06 (C3 ), 141.75 (C7),
145.98 (C17), 152.66 (C4), 160.70 (C10), 166.05 (C9). HRMS calcd
for C27H31N4O2 (MHþ), 443.2460; found, 443.2447. HPLC purity:
95%.
synthesis. Yellow solid (0.76 g, 58%); mp 195-197 °C. IR (KBr) ν cm-1
:
2975 (C-H), 1624 (CdC ethyl.), 1605 (CdC arom.), 1516 (CdC
arom.), 1252 (C-O). 1H NMR (CDCl3 300 MHz) δ ppm: 1.46 (t, 3H,
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Journal of Medicinal Chemistry
ARTICLE
J = 7.0 Hz, CH3), 4.08 (q, 2H, J = 7.0 Hz, CH2), 6.94 (d, 2H, J = 8.8 Hz,
H3,5), 7.36 (d, 1H, J = 16.5 Hz, H8), 7.60 (d, 2H, J = 8.7 Hz, H2,6), 7.81 (td,
1H, J = 7.0 Hz,J0 = 1.2 Hz, H13), 7.94 (d, 1H, J = 7.5 Hz, H12), 7.96 (td, 1H,
J = 6.7 Hz, J0 = 1.3 Hz, H14), 8.10 (d, 1H, J = 16.6 Hz, H7), 8.66 (s, 1H,
H17), 8.68 (d, 1H, J = 8.9 Hz, H15). 13C NMR (CDCl3 75 MHz) δ ppm:
14.84 (CH3), 63.59 (CH2), 108.28 (C8), 114.80 (C3,5), 123.07 (C11),
123.27 (C15), 123.69 (C16), 128.04 (C12), 128.70 (C1), 128.77 (C2,6),
130.71 (C13), 134.03 (C14), 135.74 (C7), 142.73 (C9), 147.36 (C17),
149.04 (C10), 159.87 (C4). HRMS calcd for C19H17N4O (MHþ),
317.1407; found, 317.1402. HPLC purity: 95%.
[I > 2σ(I)] = 0.0637 and wR2 [all data] = 0.1932, largest diff. peak and
hole: 0.451 and -0.237 e.Å-3
.
Crystal Data for 11a. C18H14N4O, M = 302.33, monoclinic, space
group P21/n, a = 7.9162(1) Å, b = 12.8303(2) Å, c = 14.6672(3) Å, β =
104.344(1), V = 1443.27(4) Å3, Z = 4, T = 173(2) K, 21211 reflections
collected (3557 independent, Rint = 0.0391), 209 parameters, R1 [I >
2σ(I)] = 0.0392 and wR2 [all data] = 0.1034, largest diff. peak and hole:
0.245 and -0.198 e.Å-3
.
’ ASSOCIATED CONTENT
(E)-3-(4-(20,20,20-Trifluoroethoxy)styryl)-[1,2,4]triazolo[3,4-R]phth-
alazine (13d). Compound 13d was prepared from acid 6d according to
procedure used for 13e synthesis. Yellow solid (0.29 g, 60%); mp 245-
247 °C. IR (KBr) ν cm-1: 2925 (C-H), 1640 (CdC), 1626 (CdC),
1605 (CdC arom.), 1605 (CdC arom.), 1519 (CdC arom.), 1242
(C-O), 1176 (C-F). 1H NMR (DMSO-d6, 300 MHz) δ ppm: 4.90 (q,
2H, J = 8.9 Hz, CH2), 7.19 (d, 2H, J = 8.8 Hz, H3,5), 7.52 (d, 1H, J = 166
Hz, H8), 784 (d, 2H, J = 8.8 Hz, H2,6), 8.0 (td,1H, J = 7.5 Hz, J 0 = 12 Hz,
H13), 8.05 (d, 1H, J = 16.6 Hz, H7), 8.13 (td, 1H, J = 7.4 Hz, J0 = 1.3 Hz,
H14), 8.30 (d, 1H, J = 7.5 Hz, H12), 8.59 (d, 1H, J = 7.9 Hz, H15), 9.21 (s,
1H, H17). 13C NMR (DMSO-d6, 75 MHz) δ ppm: 65.10 (q, 1C, J =
34.2 Hz, CF3), 110.04 (C8), 115.80 (C3,5), 122.63 (C15), 123.30
(C11), 123.55 (C16), 124.94 (q, J = 277.9 Hz, CH2), 129.39 (C2,6),
129.60 (C12), 130.35 (C1), 131.53 (C13), 134.57 (C7), 134.92 (C14),
142.92 (C9), 148.44 (C10), 148.91 (C17), 148.07 (C4). 19F NMR
(acetone-d6, 282 MHz) δ ppm: 74.04 (t, 3F, J = 8.7 Hz). HRMS calcd
for C19H14F3N4O (MHþ), 371.1134; found, 371.1120. HPLC pur-
ity: 95%.
S
Supporting Information. HPLC purity analysis data for
b
target compounds and crystallographic data. This material is
available free of charge via the Internet at http://pubs.acs.org.
’ AUTHOR INFORMATION
Corresponding Author
*(M.D.) Tel: (þ)33-561-175-569. Fax: (þ)33-561-175-580.
E-mail: mamadou.daffe@ipbs.fr. (M.B.) Tel: 0033(0)561556289.
Fax: 0033(0)561556011. E-mail: baltas@chimie.ups-tlse.fr.
’ ACKNOWLEDGMENT
We thank “Universitꢀe Paul Sabatier” for a postdoctoral grant
(P.D.). Thanks also to the European Community for financial
support (integrated project “New Medicines for Tuberculosis:
NM4TB 018923”). We express our sincere gratitude to Prof.
Brigitte Gicquel and Vꢀeronique Cadet-Daniel (Institut Pasteur,
Paris, France) and Catherine Pierre-Audigier (Hopital Bichat,
Paris, France) for the generous gift of INH-R strains.
3-(4-((E)-3,7-Dimethylocta-2,6-dienyloxy)styryl)-[1,2,4]triazolo-
[3,4-R]phthalazine (13f). Compound 13f was prepared from acid 6f
according to procedure used for 13e synthesis. Yellow solid (109 mg,
61%); mp 150-152 °C. IR (KBr) ν cm-1: 2969 (CdC-H), 2912 (C-
H), 2853 (C-H), 1638 (CdC), 1626 (CdC), 1604 (CdC arom.),
’ ABBREVIATIONS USED
1519 (CdC arom.), 1249 (C-O), 968 (CHdCH), 822 (C arom 1-4).
TB, tuberculosis; HIV, human immunodeficiency virus; MDR,
multidrug-resistant; XDR, extensively drug-resistant; MIC, mini-
mum inhibitory concentration; INH, isoniazid; ACP, acyl carrier
protein; DMAP, 4-N,N-dimethylaminopyridine; BSA, bis-tri-
1
0
H NMR (CDCl3, 300 MHz) δ ppm: 1.61 (s, 3H, H9 ), 1.68 (s, 3H,
0
0
0
0
H10 ), 1.76 (s, 3H, H4 ), 2.11 (m, 4H, H5 ,6 ), 4.59 (d, 2H, J = 6.6 Hz,
0
0
0
H1 ), 5.09 (t, 1H, J = 6.6 Hz, H7 ), 5.51 (t, 1H, J = 6.6 Hz, H2 ), 6.95 (d,
2H, J = 8.8 Hz, H3,5), 7.36 (d, 1H, J = 16.5 Hz, H8), 7.59 (d, 2H, J = 8.7
Hz, H2,6), 7.80 (td, 1H, J = 6.8 Hz, J0 = 1.1 Hz, H13), 7.93 (d, 1H, J = 7.6
Hz, H12), 7.95 (td, 1H, J = 7.2 Hz, J0 = 1.2 Hz, H14), 8.10 (d, 1H, J = 16.5
Hz, H7), 8.65 (s, 1H, H17), 8.68 (d, 1H, J = 7.8 Hz, H15). 13C NMR
methylsilylacetamide; EDC HCl, 1-ethyl-3-(3-dimethyllamino-
3
propyl)carbodiimide hydrochloride; HOBt, N-hydroxybenzo-
triazole; HBTU, N,N,N0,N0-tetra-methyl-O-(1H-benzotriazol-1-
yl)-uronium hexafluorophosphate; MTT, 3-(4,5-dimethylthia-
zol-2-yl)-2,5-diphenyltetrazolium bromide; SI, selectivity index;
PSA, polar surface area
0
0
0
(CDCl3 75 MHz) δ ppm: 16.71 (C9 ), 17.72 (C10 ), 25.70 (C4 ), 26.29
0
0
0
(C6 ), 39.55 (C5 ), 64.99 (C1 ), 108.34 (C8), 115.03 (C3,5), 119.24
0
0
(C2 ), 123.03 (C11), 123.22 (C15), 123.70 (C16), 123.77 (C7 ), 128.0
0
(C12), 128.69 (C1), 128.76 (C2,6), 130.64 (C13), 131.97 (C8 ), 133.97
0
(C14), 135.63 (C7), 141.49 (C3 ), 142.72 (C9), 147.29 (C17), 149.03
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