Synthesis of Natural and Unnatural Quinolones Inhibiting the Growth and Motility of Bacteria

Article abstract of DOI:10.1021/acs.jnatprod.0c00865

Synthesis of a recently discovered S-methylated quinolone natural product (1) was carried out, in addition to the production of a range of 2-substituted 4-quinolone derivatives (2-11). Two approaches were used: (i) the base-catalyzed cyclization of N-(ket

Full text of DOI:10.1021/acs.jnatprod.0c00865

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Synthesis of Natural and Unnatural Quinolones Inhibiting the
Growth and Motility of Bacteria
Jianye Li and Benjamin R. Clark*
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ABSTRACT: Synthesis of a recently discovered S-methylated
quinolone natural product (1) was carried out, in addition to the
production of a range of 2-substituted 4-quinolone derivatives (2−
11). Two approaches were used: (i) the base-catalyzed cyclization
of N-(ketoaryl)amides; (ii) attachment of the substituent to the
quinolone core via a Suzuki−Miyaura cross-coupling. Also
produced were a small suite of related 2(1H)-quinolones (12−
19). The synthesized compounds were assessed for their
antimicrobial properties. The alkene-substituted 4-quinolone 8
significantly inhibited the growth of a Pseudomonas aeruginosa
strain, and both 4-quinolones and 2(1H)-quinolones were capable
of inhibiting the swarming behavior of Bacillus subtilis.
derivatives could affect the swarming behavior and biofilm
he 2-alkyl-4-quinolones are a group of microbial natural
Tproducts, the best known of which are the Pseudomonas formation of Bacillus species.16,17
quinolone signal (PQS)1 and its biosynthetic precursor, 4-
hydroxy-2-heptylquinoline (HHQ).2 These metabolites play a
role in quorum sensing of Pseudomonas aeruginosa,3,4 which
influences biofilm formation and virulence factor production;
inhibition of quinolone production can thus be used as a
strategy for the control of pathogenic Pseudomonas strains.5,6 A
related suite of compounds are produced in some Burkholderia
species, where they fulfill a similar role.7 Other members of the
structure class have been reported to possess antialgal,8
antimalarial, and cytotoxic properties.9 Structural variation in
the majority of the naturally occurring 2-alkyl-4-quinolones
centers on the substituent at the 2-position, which usually
consists of saturated or unsaturated alkyl chains.
In order to expand the chemical space available for biological
investigation, many researchers have taken a synthetic
approach.10 Fluorinated 2-phenyl-4-quinolones have displayed
potent cytotoxic properties, acting as tubulin inhibitors,11 while
2-phenyl-4-quinolone itself has been shown to induce G2/M
phase arrest in a leukemia cell line.12 In 2007, a two-step
synthetic procedure for the synthesis of quinolones was
described, using a Cu-catalyzed Camps cyclization to produce
a range of 2-substituted 4-quinolones.13 A 2016 report
described the synthesis of a range of HHQ and PQS
derivatives in which the heteroatoms in the quinolone ring
were replaced,14 one of which displayed significant inhibitory
properties against P. aeruginosa elastase, which is a key
virulence factor for the microorganism and a potential drug
target.15 Alkylquinolones affect not only the Pseudomonas
genus, however: reports in 2012 and 2015 revealed that HHQ
Quinolones are also known for their antibiotic properties.
Best known of these are the synthetic fluoroquinolones,18
which possesses significantly different substitution patterns
from the naturally occurring 2-alkyl-4-quinolones, but the
latter class and its derivatives have also been reported to have
antimicrobial properties. 3-Thiocyanate- and selenocyanate-
substituted 4-arylquinolones have been shown to possess
moderate antimicrobial properties against both Gram-positive
and Gram-negative bacteria.19 In a report by Salvaggio and co-
workers, a small suite of quinolone natural products was
synthesized using a Suzuki−Miyaura coupling, several
members of which were found to retard the growth of E. coli
and S. aureus in vitro.20 The same method has been used to
produce Burkholderia-derived quinolones, which possess
antimicrobial properties against Gram-positive bacteria.21
In a recent report, we described the isolation of several 4-
quinolone natural products from a P. aeruginosa strain, several
of which possessed unusual side chains at the 2-position.22
These compounds possessed weak cytotoxic activity, but
insufficient material was isolated for further biological testing;
they thus presented attractive synthetic targets. Of chief
interest was an unprecedented compound containing a 2-
Received: August 3, 2020
© XXXX American Chemical Society and
American Society of Pharmacognosy
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methylthiovinyl group (1). We describe here the first synthesis
of this unusual natural product, along with a range of 2-
substituted 4-quinolone derivatives (2−11), two of which are
also recently discovered natural products (6, 8). In addition, a
small suite of related quinolin-2(1H)-ones (12−19) were
produced, and the antimicrobial properties of all synthesized
compounds were assessed.
Scheme 1. Retrosynthetic Analysis for Synthesis of 2-
Substituted Quinolones
RESULTS AND DISCUSSION
■
Retrosynthetic Analysis. 2-Substituted quinolones in
which the side chains contain double bonds conjugated to
the core nucleus can be synthesized by the base-promoted
cyclization of N-(ketoaryl)amides.13 The N-(ketoaryl)amides
can be obtained by condensation of 2′-aminoacetophenone
(20) with the corresponding carboxylic acid or acyl chloride
starting materials, which are either commercially available or
easy to synthesize. Another method is by attaching the
substituent at the 2-position of the quinolone core in the final
step, via a Suzuki−Miyaura cross-coupling. The halogenated
quinolone derivative 21 can be easily obtained from the ester
22, which in turn is derived from aniline (23) and
dimethylacetylene dicarboxylate (24) by the Conrad−Limpach
method, followed by reduction and chlorination.20
With a synthetic version of the natural product in hand and a
viable methodology, our attention turned to the production of
derivatives. Using sodium ethanethiolate, the amide 28 was
formed from 25 as the major product, with a small amount of
the bis-adduct 29 formed as a side product. Cyclization of 28
under basic conditions yielded the ethyl sulfide-substituted
quinolone 3; the Z-isomer was not detected. Oxygenated
derivatives were also produced, with the conjugated amides 30
and 31 formed by condensation of 20 with the corresponding
carbonyl compounds (Scheme 3). The amides were sub-
sequently cyclized under basic conditions to yield the methyl
(4) and ethyl ether-substituted quinolones (5). Yields for the
cyclization of the ethers were much lower than those for the
sulfides, with significant amounts of unknown side products
formed.
The structure of the side product from the cyclization of 31
was determined by !D and 2D NMR experiments. In the
proton NMR spectrum, signals corresponding to a 1,2-
disubsituted aromatic ring were present, in addition to singlet
resonances consistent with the presence of an aromatic methyl
group (δH 2.78, 3H), two identical ethoxy groups (δH 1.27, t,
6H; 3.62, dq, 2H; 3.87, dq, 2H), and a highly deshielded
methine (δH 6.27, 1H). HRMS analysis revealed a sodium
adduct ion consistent with a molecular formula of C15H19NO3.
In combination with 2D NMR analysis (Supporting
Information), this was sufficient to establish the structure as
shown (17). The structure of the side product from the
methoxylated amide 30 was established in a similar manner
(16). In order to broaden the structural diversity of these
unusual side products for later biological investigations, two
additional analogues were produced by base-catalyzed
cyclization of the sulfur-containing amides 27 and 29,
synthesized previously, to yield the quinolones 18 and 19.
Carboxylic acids lacking double bonds were readily
condensed with 2′-aminoacetophenone using EDCI as the
condensing agent (Scheme 4).24 In this manner, the amides 32
and 33 were produced by condensation of the appropriate
Synthesis of Quinolones. Initial attempts at the synthesis
of the natural product 1 followed the former strategy: 3-
methylthioacrylic acid was synthesized by the addition of
NaSMe to freshly distilled propiolic acid to yield a mixture of
cis−trans isomers (trans:cis 40:1).23 Condensation with 2′-
aminoacetophenone (20) using N,N′-dicyclohexylcarbodii-
mide (DCC) and 1-ethyl-3-(3-(dimethylamino)propyl)-
carbodiimide hydrochloride (EDCI) was attempted, but the
reaction did not proceed efficiently, while the reaction using
the acid chloride yielded a complex mixture. Thus, an
alternative approach was used: first 2′-aminoacetophenone
was condensed with propiolic acid to synthesize the
propiolamide derivative 25, using DCC as the coupling agent
at 60 °C in tetrahydrofuran (THF), with a yield of 60%. The
key intermediate 26 was synthesized by the addition of NaSMe
to the alkynyl group in good yield (78%). However, a
significant amount of a side product, the bis(methylthio)-
propanamide 27, was also formed, resulting from addition of
two NaSMe units to the triple bond. Finally, the target
quinolone 1 was achieved by NaOH-promoted cyclization of
26 at 110 °C in dioxane for 1 h (Scheme 2), along with the Z-
isomer 2, in a 5:1 ratio. Spectroscopic data for the target
quinolone were identical to those for the natural product.22
B
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Scheme 2. Synthesis of the Natural Product 1 and Related Sulfur-Containing Quinolones
Scheme 3. Synthesis of Quinolones with Oxygen-Containing Side Chains
Scheme 4. Synthesis of 2(1H)-Quinolones 12−15
carboxylic acid with 2′-aminoacetophenone in high yield.
Addition of NH(CH3)2 and NaSCH2CH3 to the amide 34
yielded further amides 35 and 36. All four amides were
cyclized under basic conditions; unfortunately, the desired
cyclization did not take place, leading to the formation of
2(1H)-quinolones (12−15) instead of the desired quinolin-4-
ones (Scheme 4). Nevertheless, some 2(1H)-quinolones have
been shown to possess significant activity against microbial
pathogens,25 and as such the compounds were retained for
testing.
Thus, an alternative synthetic approach was required for the
synthesis of the remaining quinolones. The 2-(chloromethyl)-
quinolone scaffold was formed by the Conrad−Limpach
approach20 in high yield (Scheme 5). Aniline (23) was
condensed with dimethylacetylene dicarboxylate (24) in
MeOH at reflux to furnish diester 37, which underwent
polyphosphoric acid (PPA)-promoted intramolecular Friedel−
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Scheme 5. Synthesis of 4-Quinolones 6−11 via Suzuki−Miyaura Cross-Coupling
a
Table 1. Zone of Inhibition (mm) of Active Quinolones against Test Bacteria in Well Diffusion Assays
Staphylococcus aureus ATCC
Staphylococcus epidermidis ATCC
Pseudomonas aeruginosa ATCC Bacillus subtilis subsp. spizizenii ATCC
compound
12600
14990
15692
6633
1
3
7
8
−
13*
−
9*
−
39
n/t
n/t
−
−
−
24*
−
29
n/t
n/t
−
−
29*
35*
36*
n/t
34
23*
−
−
9*
−
n/t
n/t
19
10
ampicillin
ciprofloxacin
vancomycin
n/t
a
− = no activity; n/t = not tested; * = inhibition halo was hazy/indistinct.
Crafts acylation to obtain 1,4-dihydroquinoline-2-carboxylate
22. The subsequent reduction of the ester in the presence of
sodium borohydride afforded the alcohol 38 in good yield.
Substitution of the hydroxy group by chlorination with thionyl
chloride led to the formation of the halide-substituted
quinolone core 21 in good yield.
In order to produce the quinolone targets, a Suzuki−
Miyaura coupling reaction with boronic acid derivatives was
used. Commercially available pinacolboronates and boronic
acids were reacted with the halide-substituted quinolone 21
under the catalytic action of a dichloro(1,1′-bis-
(diphenylphosphanyl)ferrocene)palladium(II) complex, lead-
ing to the formation of desired products (6−8) in good yields.
Spectroscopic data for 6 and 8 matched well with literature
values for the natural products.22 The corresponding 1-methyl-
substituted quinolones (9−11) were synthesized using the
same methodology from N-methylaniline (39).
Biological Testing. All quinolones were subjected to
preliminary antimicrobial screening using well-diffusion assays
against a range of eight Gram-positive and Gram-negative test
microbes (Supporting Information Table S1). A small number
of quinolones displayed activity in these assays, with 1
displaying activity against B. subtilis and 3 against S. aureus,
while 7 and 10 showed significant inhibition halos against P.
aeruginosa (Table 1). Most notable though was the unsaturated
side chain quinolone 8, which displayed activity against four of
the tested strains. Results for the active compounds showed a
distinctive hazy zone of inhibition (Supporting Information
Figure S1). Such an effect can be associated with overlong
growth times or an excessively concentrated inoculum, but the
results were reproducible even with shorter growth times and a
more dilute inoculum. Thus, this might represent partial
growth inhibition, rather than a bactericidal effect. PQS itself is
known to inhibit the growth of P. aeruginosa, so it was possible
these compounds were exhibiting a similar effect.26
In order to further investigate this effect, selected quinolones
were assessed for their effects in a growth inhibition assay
against P. aeruginosa. While the majority of the quinolones
tested did not act as straightforward bacteriocides, with
residual growth observed even at high concentrations, several
of them nevertheless significantly inhibited the growth of P.
aeruginosa. (Figure 1). The natural product 8 showed strong
growth inhibition at all tested concentrations, while the
remainder of the 4-quinolones delayed the growth of P.
aeruginosa by 37−58% at concentrations between 2.1 and 33.4
μg/mL. Compound 8 has been shown to possess antimicrobial
properties in a prior report, against the fish pathogen
Tenacibaculum maritimum.27 In contrast, the 2(1H)-quinolones
18 and 19 had minimal activity.
In order to further assess the effects of the compounds over
time, time−kill experiments were conducted on compound 8,
as it was the only compound giving a measurable MIC value
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aeruginosa.20 In contrast, a sulfur-containing derivative of PQS
did not possess any significant growth inhibitory properties,14
highlighting the very specific substitution patterns required for
this activity.
Previous studies have shown that alkylquinolones can affect
the swarming motility in a number of microbial species.
Swarming motility is the movement of bacteria across a
semisolid surface, a process that is involved in both the
virulence and antibiotic resistance of many bacterial species.28
PQS itself is a potent inhibitor of swarming behavior in both P.
aeruginosa and other Gram-negative and -positive bacteria,
while HHQ exhibits more moderate effects.29,30 Prior synthetic
studies have shown that the substitution at the 3-position is
also crucial for activity, with a suite of C-3 halogenated
derivatives lacking activity against B. subtilis swarming
motility.16 In a subsequent study, a range of derivatives
substituted at different points around the aromatic ring were
produced, with 6-substituted analogues displaying potent
activity in inhibiting swarming behavior in B. atrophaeus.17
However, the effect of changing the C-2 substituent has not
been investigated in detail to date. Thus, the activity of the
quinolones 1−19 against the swarming behavior of B. subtilis
was assessed. Of the 4-quinolones, 6, 8, 10, and 11 possessed
comparable swarming inhibition activity to HHQ, while the
aromatic sulfide derivative 7 had significantly enhanced activity
(Figure 3). 4-Quinolones containing a shorter side chain (1−
5) had weak antiswarming activity; those with a longer chain
(8) or a bulky aromatic substituent (6, 7) were more active.
The effect of N-methylation was less clear: N-methylated
derivatives 9 and 10 lost activity compared to their
nonmethylated analogues, though compound 11 did not. For
the 2-quinolones, all of the acetal and thioacetal derivatives
(16−19) possessed weak activity, while ether- and sulfide-
substituted quinolones 12, 13, and 15 were of similar or
greater potency to HHQ. Compound 14 possessed signifi-
cantly lower activity than the former three compounds, which
might be attributed to the presence of the ionizable amine
group in the side chain of 14.
Figure 1. Inhibition of selected compounds against P. aeruginosa
ATCC 15692, with ciprofloxacin (Cip) as the positive control. For
each compound, the six columns from left to right represent the
extent of inhibition at final concentrations of 1.1 (yellow), 2.1 (dark
green), 4.2 (pale green), 8.4 (pink), 16.7 (blue), and 33.4 (deep
purple) μg/mL, respectively.
(33 μg/mL). At 1/4 the MIC value, significant inhibition of
growth was observed, with a small drop in CFU values
observed after 2 h and very little growth observed out to 24 h
(Figure 2). At the MIC value, similar but more dramatic effects
were observed, with LogCFU values decreasing steadily until
10 h, before stabilizing. Even at very high concentrations (4 ×
MIC), while a rapid decrease in CFU values was observed
initially, viable bacteria were still detectable after 24 h, in
contrast to the positive control (ciprofloxacin). Other
quinolones have been shown to possess similar growth
inhibitory effects: several Pseudonocardia-derived isoprenylated
4-quinolones have shown a similar effect against E. coli and S.
aureus strains, where a significant delay in the onset of growth
was observed, though they did not affect the growth of P.
Overall, these studies further elucidate the numerous ways in
which the alkylquinolones can affect the growth and mobility
Figure 2. Time−kill curves of compound 8 (a) in comparison with ciprofloxacin (b) showing colony forming units (CFUs) for P. aeruginosa ATCC
15692 at different time points. P. aeruginosa were supplemented with compound 8 and ciprofloxacin at 0.25 × MIC (red line), at the MIC (MIC,
blue line), and at four times the MIC (4 × MIC, purple line) or DMSO (vehicle) (black line). The MIC for 8 was 33 μg/mL, while the MIC value
for ciprofloxacin was 4 μg/mL.
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7.57 (t, J = 7.9 Hz, 1H), 7.92 (dd, J = 8.0, 1.5 Hz, 1H), 8.66 (d, J =
8.2 Hz, 1H), 12.12 (s, 1H); ESIMS m/z 188.07 [M + H]+.
(E)-N-(2-Acetylphenyl)-3-(methylthio)acrylamide (26) and N-(2-
Acetylphenyl)-3,3-bis(methylthio)propanamide (27). A mixture of
compound 25 (100.0 mg, 0.53 mmol) and 20% sodium methanethiol
solution (39.3 mg, 0.56 mmol) in THF was heated at 40 °C for 4 h.
The mixture was evaporated under reduced pressure, transferred into
a separatory funnel, and extracted with H2O (30 mL) and EtOAc (3
× 20 mL), and the combined organic layers were dried over
anhydrous MgSO4 and evaporated under reduced pressure. The
residue was purified by silica gel flash chromatography (hexanes−
EtOAc, 25:1 to 5:1) to give amide 26 (98.2 mg, 79%) as a pale yellow
solid and 27 (25.0 mg, 16%) as a white gum. When the amount of
sodium methanethiol was doubled and the temperature increased to
60 °C, the yield of compound 27 was 65%.
Compound 26: 1H NMR (400 MHz, methanol-d4) δ 2.43 (s, 3H),
2.65 (s, 3H), 5.91 (d, J = 14.6 Hz, 1H), 7.18 (t, J = 8.7 Hz, 1H), 7.55
(t, J = 8.6 Hz, 1H), 7.76 (d, J = 14.6 Hz, 1H), 8.02 (dd, J = 8.0, 1.3
Hz, 1 H), 8.57 (d, J = 8.4 Hz, 1H); 13C NMR (100 MHz, methanol-
d4) δ 14.5, 28.7, 117.4, 121.9, 124.0, 124.3, 133.1, 135.6, 141.5, 146.4,
165.3, 204.6; HRESIMS m/z 258.0549 [M + Na]+ (calcd for
C12H13NO2S, 258.0559).
Compound 27: 1H NMR (600 MHz, CDCl3) δ 2.17 (s, 6H), 2.67
(s, 3H), 2.88 (d, J = 7.5 Hz, 2H), 4.27 (t, J = 7.5 Hz, 1H), 7.13 (t, J =
7.5 Hz, 1H), 7.56 (t, J = 8.1 Hz, 1H), 7.90 (dd, J = 8.0, 1.4 Hz, 1H),
8.76 (d, J = 8.1 Hz, 1H), 11.83 (s, 1 H); 13C NMR (150 MHz,
CDCl3) δ 13.2, 28.7, 44.8, 50.2, 121.1, 122.0, 122.8, 131.8, 135.4,
140.9, 168.6, 203.1; HRESIMS m/z 306.0592 [M + Na]+ (calcd for
C13H17NO2S2, 306.0593).
Figure 3. Effect of quinolones 1−19 (10 μM) on swarming motility
of B. subtilis on BHI agar. (a) Visual representation of the data
showing representative plates; (b) average diameter of swarming zone
from three replicates.
(E)-N-(2-Acetylphenyl)-3-(ethylthio)acrylamide (28) and N-(2-
Acetylphenyl)-3,3-bis(ethylthio)propanamide (29). The procedure
was the same as for the synthesis of compounds 26 and 27, using
sodium ethanethiolate (47.0 mg, 0.56 mmol). Compound 28 (103.0
mg, 78%) as a pale yellow solid and compound 29 (28.2 mg, 17%)
were isolated. When the amount of sodium ethanethiolate was
doubled and the solution was heated at 60 °C, the yield of compound
29 was 67%.
of microbial species. The results suggest that further
investigation of aromatic-substituted 4-quinolones may be
valuable in the discovery of compounds interfering with
bacterial growth and swarming behavior, while the serendip-
itously obtained 2(1H)-quinolones provide an additional
avenue of research.
Compound 28: 1H NMR (600 MHz, CDCl3) δ 1.37 (t, J = 7.4 Hz,
3H), 2.66, (s, 3H), 2.86 (q, J = 7.4 Hz, 2H), 5.92 (d, J = 14.8 Hz,
1H), 7.09 (t, J = 7.5 Hz, 1H), 7.54 (t, J = 7.8 Hz, 1H), 7.72 (d, J =
14.8 Hz, 1H), 7.88 (dd, J = 8.0, 1.5 Hz, 1H), 8.82 (dd, J = 8.5, 1.0 Hz,
1H), 11.73 (s, 1H); 13C NMR (150 MHz, CDCl3) δ 14.0, 26.5, 28.7,
117.8, 121.0, 121.8, 122.2, 131.8, 135.3, 141.7, 144.3, 163.6, 203.0;
ESIMS m/z 250.08 [M + H]+.
EXPERIMENTAL SECTION
■
General Experimental Procedures. NMR spectra were
recorded on a Bruker Avance III 400 spectrometer and a Bruker
Avance III 600 spectrometer with tetramethylsilane as an internal
reference. HRESIMS spectra were obtained using a Thermo Fisher Q
Exactive HF mass spectrometer or an Agilent 6230 HPLC system
with a Bruker microTOF-QII mass spectrometer. Low-resolution MS
spectra were acquired using an Agilent 6430 Triple-Quadrupole
instrument. HPLC analysis was performed on an Agilent 1260 series
HPLC system equipped with a G1311B quaternary pump, a G1315D
photodiode array detector, a G1316C thermostated column compart-
ment, and a G1329B autosampler, using a reversed-phase ZORBAX
SB-C18 column (3.5 μm, 4.6 × 150 mm; Agilent Technologies).
Semipreparative HPLC separation was carried out on a Shimadzu LC-
20AR series HPLC instrument equipped with a UV detector and a
reversed-phase C18 column (ZORBAX SB-C18, 5 μm, 9.4 × 250
mm). Reagents and solvents used in this study were commercially
available (Jiangtian, Heowns, Concord Tianjin). All reactions were
monitored by TLC using commercial silica gel plates.
Compound 29: 1H NMR (600 MHz, CDCl3) δ 1.28 (t, J = 7.4 Hz,
6H), 2.66, (s, 3H), 2.62−2.78 (m, 4H), 2.91 (d, J = 7.4 Hz, 2H), 4.40
(t, J = 7.4 Hz, 1H), 7.13 (t, J = 7.6 Hz, 1H), 7.55 (t, J = 7.9 Hz, 1H),
7.90 (dd, J = 8.0, 1.5 Hz, 1H), 8.76 (dd, J = 8.5, 0.9 Hz, 1H), 11.82 (s,
1H); 13C NMR (150 MHz, CDCl3) δ 14.6, 24.9, 28.7, 46.1, 46.7,
121.0, 122.0, 122.7, 131.7, 135.3, 140.9, 168.8, 203.0; ESIMS m/z
312.16 [M + H]+.
General Procedure A for the Synthesis of Amides. A mixture
of 1-(2-aminophenyl)ethanone 20 (0.5 g, 3.7 mmol), acid (4.1
mmol), EDCI (0.85 g, 4.44 mmol), 4-dimethylaminopyridine
(DMAP) (45.0 mg, 0.37 mmol), and triethylamine (TEA) (0.77
mL, 5.6 mmol) in CH2Cl2 was stirred overnight at room temperature
(rt). The mixture was transferred into a separatory funnel and
extracted with H2O (30 mL) and CH2Cl2 (3 × 20 mL). The
combined organic layers were dried over anhydrous MgSO4 and
evaporated under reduced pressure. The residue was purified by silica
gel flash chromatography (hexanes−EtOAc, 25:1 to 5:1).
N-(2-Acetylphenyl)propiolamide (25). A mixture of 1-(2-
aminophenyl)ethanone 20 (0.5 g, 3.7 mmol), propiolic acid (0.28
g, 4.1 mmol), and DCC (1.14 g, 5.55 mmol) in THF was heated at 60
°C for 12 h. The reaction mixture was evaporated under reduced
pressure, transferred into a separatory funnel, and extracted with H2O
(70 mL) and EtOAc (3 × 50 mL), and the combined organic layers
were dried over anhydrous MgSO4 and evaporated under reduced
pressure. The residue was purified by silica gel flash chromatography
(hexanes−EtOAc, 25:1 to 5:1) to give the amide 25 as a yellow solid
(0.40 g, 55%). A small amount of an N-acylurea impurity was present,
but the material was used directly for the next reaction. 1H NMR (400
MHz, CDCl3) δ 2.68 (s, 3H), 2.95, (s, 1H), 7.18 (t, J = 7.6 Hz, 1H),
(E)-N-(2-Acetylphenyl)-3-methoxyacrylamide (30). General pro-
cedure A using (E)-3-methoxyacrylic acid (0.42 g, 4.1 mmol) yielded
1
compound 30 (0.44 g, 54%) as a pale yellow solid: H NMR (400
MHz, CDCl3) δ 2.66 (s, 3H), 3.73, (s, 3H), 5.38 (d, J = 12.2 Hz, 1H),
7.08 (t, J = 7.8, 1H), 7.54 (t, J = 8.1 Hz, 1H), 7.69 (d, J = 12.2 Hz,
1H), 7.88 (dd, J = 8.0, 1.4 Hz, 1H), 8.81 (dd, J = 8.6, 1.2 Hz, 1H),
11.65 (s, 1H); 13C NMR (100 MHz, CDCl3) δ 28.8, 57.6, 99.8, 121.0,
121.6, 122.0, 131.8, 135.3, 141.8, 161.9, 165.9, 203.0; HRESIMS m/z
242.0794 [M + Na]+ (calcd for C12H13NO3, 242.0788).
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(E)-N-(2-Acetylphenyl)-3-ethoxyacrylamide (31). A solution of 1-
(2-aminophenyl)ethanone 20 (0.2 g, 1.5 mmol) and TEA (0.27 mL,
1.9 mmol) in THF was added dropwise to a solution of (E)-3-
ethoxyacryloyl chloride (0.22 g, 1.6 mmol) at 0 °C. The reaction
mixture was warmed to rt and stirred for 4 h. Then the mixture was
evaporated under reduced pressure, transferred into a separatory
funnel, and extracted with H2O (50 mL) and EtOAc (3 × 30 mL),
and the combined organic layers were dried over anhydrous MgSO4
and evaporated under reduced pressure to give amide 31 (342.1 mg,
98%) as a pale yellow solid: 1H NMR (400 MHz, CDCl3) δ 1.36 (t, J
= 7.1 Hz, 3H), 2.66 (s, 3H), 3.95 (q, J = 7.1 Hz, 2H), 5.39 (d, J = 12.2
Hz, 1H), 7.07 (t, J = 7.6, 1H), 7.53 (t, J = 7.9 Hz, 1H), 7.65 (d, J =
12.2 Hz, 1H), 7.88 (dd, J = 8.0, 1.4 Hz, 1H), 8.81 (dd, J = 8.6, 1.2 Hz,
1H), 11.61 (s, 1H); 13C NMR (100 MHz, CDCl3) δ 14.7, 28.7, 67.1,
100.3, 121.0, 121.6, 121.9, 131.8, 135.3, 141.9, 161.1, 166.2, 203.0;
HRESIMS m/z 256.0960 [M + Na]+ (calcd for C13H15NO3,
256.0944).
20 mL), and the combined organic layers were dried over anhydrous
MgSO4 and evaporated under reduced pressure. The residue was
purified by silica gel flash chromatography (hexanes−EtOAc, 25:1 to
1
5:1) to give amide 36 (127.7 mg, 96%) as a pale yellow solid: H
NMR (400 MHz, CDCl3) δ 1.23 (t, J = 7.4 Hz, 3H), 2.55 (q, J = 7.4
Hz, 2H), 2.61 (s, 3H), 2.67 (t, J = 7.4 Hz, 2H), 2.87 (t, J = 7.3 Hz,
2H), 7.06 (t, J = 7.6, 1H), 7.49 (t, J = 7.9 Hz, 1H), 7.84 (d, J = 7.9 Hz,
1H), 8.70 (d, J = 8.5 1H), 11.75 (s, 1H); 13C NMR (100 MHz,
CDCl3) δ 14.7, 26.0, 26.9, 28.6, 38.8, 120.7, 121.6, 122.4, 131.6,
135.1, 140.8, 170.5, 202.8; ESIMS m/z 274.08 [M + H]+.
General Procedure B for Base-Promoted Cyclization. A
resealable oven-dried test tube with a stir bar was charged with N-
(ketoaryl)amide (1 equiv) and crushed NaOH (3 equiv). Anhydrous
1,4-dioxane was added via syringe and the flask charged with argon
gas. Then the reaction was placed in a preheated oil bath at 110 °C,
stirred for 1−2 h, and then cooled to rt. The reaction mixture was
then dissolved in MeOH, transferred to a round-bottom flask, and
evaporated under reduced pressure. The residue was purified by silica
gel flash chromatography (CH2Cl2−MeOH, 100:0 to 10:1).
(E)- and (Z)-2-(2-(Methylthio)vinyl)quinolin-4(1H)-one (1 and 2).
Following general procedure B with compound 26 (36 mg, 0.15
mmol) as starting material produced an organic residue that was
purified by silica gel flash chromatography (CH2Cl2−MeOH, 100:0 to
10:1) and semipreparative HPLC with 15% MeCN−H2O to 50%
MeCN−H2O to yield compound 1 as a white solid (15 mg, 45%) and
2 as a white solid (3 mg, 10%).
N-(2-Acetylphenyl)-3-(methylthio)propanamide (32). Following
general procedure A, using 3-(methylthio)propionic acid (0.49 g, 4.1
mmol) as a starting material, compound 32 (0.81 g, 92%) was
1
obtained as a pale yellow solid: H NMR (400 MHz, CDCl3) δ 2.07
(s, 3H), 2.56 (s, 3H), 2.65 (t, J = 7.0 Hz, 2H), 2.80 (t, J = 7.0 Hz,
1H), 7.02 (t, J = 7.7, 1H), 7.44 (t, J = 7.7 Hz, 1H), 7.80 (d, J = 7.7 Hz,
1H), 8.65 (d, J = 8.0, Hz, 1H), 11.72 (s, 1H); 13C NMR (100 MHz,
CDCl3) δ 15.5, 28.4, 29.4, 38.2, 120.5, 121.5, 122.3, 131.5, 135.0,
140.6, 170.3, 202.7; HRESIMS m/z 260.0708 [M + Na]+ (calcd for
C12H15NO2S, 260.0716).
1
Compound 1: H NMR (400 MHz, methanol-d4) δ 2.47 (s, 3H),
N-(2-Acetylphenyl)-3-methoxypropanamide (33). Following gen-
eral procedure A, using 3-(methoxy)propionic acid (0.42 g, 4.0
mmol) as a starting material, compound 33 (0.77 g, 95%) was
6.18 (d, J = 15.5 Hz, 1H), 6.37 (s, 1H), 7.36 (t, J = 7.4 Hz, 1H), 7.57
(d, J = 8.3 Hz, 1H), 7.66 (t, J = 7.7 Hz, 1H), 7.68 (d, J = 15.5 Hz, 1
H), 8.17 (d, J = 8.3 Hz, 1H); 13C NMR (100 MHz, methanol-d4) δ
14.4, 105.5, 116.4, 119.1, 125.1, 125.8, 125.9, 133.5, 139.6, 141.4,
149.9, 180.3; HRESIMS m/z 218.0634 [M + H]+ (calcd for
C12H11NOS, 218.0634).
1
obtained as a pale yellow solid: H NMR (400 MHz, CDCl3) δ 2.63
(s, 3H), 2.68 (t, J = 6.0 Hz, 2H), 3.39 (s, 3H), 3.75 (t, J = 6.0 Hz,
2H), 7.08 (m, 1H), 7.51 (m, 1H), 7.85 (t, J = 6.6 Hz, 1H), 8.74 (dd, J
= 8.5, 4.8 Hz, 1H), 11.74 (s, 1H); 13C NMR (100 MHz, CDCl3) δ
28.7, 39.2, 58.9, 68.3, 121.0, 122.1, 122.5, 131.6, 135.1, 140.8, 170.5,
202.6; ESIMS m/z 222.12 [M + H]+.
1
Compound 2: H NMR (400 MHz, methanol-d4) δ 2.55 (s, 3H),
6.31 (d, J = 11.2 Hz, 1H), 6.74 (s, 1H), 7.08 (d, J = 11.2 Hz, 1H),
7.39 (t, J = 7.6 Hz, 1H), 7.55 (d, J = 8.4 Hz, 1H), 7.69 (ddd, J = 8.4,
7.0, 1.4 Hz, 1 H), 8.20 (d, J = 8.3 Hz, 1H); HRESIMS m/z 218.0634
[M + H]+ (calcd for C12H11NOS, 218.0634).
(E)-2-(2-(Ethylthio)vinyl)quinolin-4(1H)-one (3). Following gen-
eral procedure B with compound 28 (44 mg, 0.18 mmol) as starting
material, compound 3 was obtained as a pale yellow solid (18 mg,
44%): 1H NMR (600 MHz, methanol-d4) δ 1.38 (t, J = 7.4, 3H), 2.94
(q, J = 7.5, 2H), 6.26 (d, J = 15.7 Hz, 1H), 6.34 (s, 1H), 7.34 (t, J =
7.4 Hz, 1H), 7.56 (d, J = 8.3 Hz, 1H), 7.61 (d, J = 15.8 Hz, 1 H), 7.64
(t, J = 7.8 Hz, 1H), 8.16 (d, J = 8.3 Hz, 1H); 13C NMR (150 MHz,
methanol-d4) δ 14.6, 27.0, 105.5, 117.5, 119.1, 125.0, 125.8, 125.9,
133.4, 139.2, 141.4, 149.9, 180.2; HRESIMS m/z 232.0790 [M + H]+
(calcd for C13H13NOS, 232.0791).
3-(Bis(methylthio)methyl)-4-methylquinolin-2(1H)-one (18). Fol-
lowing general procedure B, with compound 27 (35 mg, 0.12 mmol)
as starting material, compound 18 was obtained as a pale yellow solid
(24 mg, 74%): 1H NMR (600 MHz, DMSO-d6) δ 2.20 (s, 6H), 2.68
(s, 3H), 5.78 (s, 1H), 7.21 (t, J = 7.3 Hz, 1H), 7.30 (d, J = 8.1 Hz,
1H), 7.50 (t, J = 7.5 Hz, 1H), 7.77 (d, J = 7.9 Hz, 1 H), 11.89 (s, 1H);
13C NMR (150 MHz, DMSO-d6) δ 16.6, 28.9, 48.2, 115.2, 120.1,
N-(2-Acetylphenyl)acrylamide (34). Using the same procedure as
for the synthesis of compound 31, using 1-(2-aminophenyl)ethanone
20 (0.16 g, 1.2 mmol) and acryloyl chloride (0.12 g, 1.3 mmol)
yielded the amide 34 (227.6 mg, 98%) as a white semisolid solid: 1H
NMR analysis revealed the presence of minor impurities remaining;
the compound was used without additional purification in subsequent
reactions. 1H NMR (600 MHz, CDCl3) δ 2.68 (s, 3H), 5.79 (dd, J =
10.3, 1.0 Hz, 1H), 6.32 (dd, J = 17.1, 10.3 Hz, 1H), 6.43 (dd, J = 17.1,
1.0 1H), 7.14 (t, J = 7.6 Hz, 1H), 7.58 (t, J = 7.9 Hz, 1H), 7.92 (dd, J
= 8.0, 1.5 Hz, 1H), 8.85 (dd, J = 8.5, 1.0 Hz, 1H), 11.98 (s, 1H); 13C
NMR (150 MHz, CDCl3) δ 28.6, 120.9, 121.9, 122.7, 127.4, 131.8,
132.6, 135.2, 141.1, 164.5, 203.0; HRESIMS m/z 212.0695 [M +
Na]+ (calcd for C11H11NO2, 212.0682).
N-(2-Acetylphenyl)-3-(dimethylamino)propanamide (35). A mix-
ture of compound 34 (100.0 mg, 0.53 mmol) and a 40%
dimethylamine aqueous solution (112.5 mg, 1.0 mmol) in THF was
heated at 60 °C for 4 h. The mixture was evaporated under reduced
pressure and transferred into a separatory funnel and extracted with
water (30 mL) and EtOAc (3 × 20 mL), and the combined organic
layers were dried over anhydrous MgSO4 and evaporated under
reduced pressure. The residue was purified by silica gel flash
chromatography (hexanes−EtOAc, 25:1 to 5:1) to give amide 35
(117.8 mg, 95%) as a pale yellow solid: 1H NMR (400 MHz, CDCl3)
δ 2.23 (s, 6H), 2.53 (t, J = 6.9 Hz, 2H), 2.57 (s, 3H), 2.66 (t, J = 6.9
Hz, 2H), 7.02 (t, J = 7.6, 1H), 7.44 (t, J = 7.9 Hz, 1H), 7.79 (dd, J =
8.0, 1.4 Hz, 1H), 8.64 (dd, J = 8.5, 0.8 Hz, 1H), 11.73 (s, 1H); 13C
NMR (100 MHz, CDCl3) δ 28.6, 36.5, 46.1, 54.9, 121.0, 122.3, 122.3,
131.4, 134.8, 140.5, 171.1, 202.4; HRESIMS m/z 235.1418 [M + H]+
(calcd for C13H19N2O2, 235.1441).
121.9, 124.7, 129.6, 130.3, 137.1, 145.9, 160.4; HRESIMS m/z
288.0489 [M + Na]+ (calcd for C13H15NOS2, 288.0487).
3-(Bis(ethylthio)methyl)-4-methylquinolin-2(1H)-one (19). Fol-
lowing general procedure B, with compound 29 (18 mg, 0.06
mmol) as starting material, compound 19 was obtained as a pale
yellow solid (12 mg, 73%): 1H NMR (400 MHz, CDCl3) δ 1.34 (t, J
= 7.4 Hz, 6H), 2.74 (q, J = 7.4 Hz, 4H), 2.81 (s, 3H), 6.16 (s, 1H),
7.24 (t, J = 7.4, 1H), 7.37 (d, J = 8.2 Hz, 1H), 7.49 (d, J = 7.8 Hz,
1H), 7.76 (d, J = 8.5 1H), 12.28 (s, 1H); 13C NMR (100 MHz,
CDCl3) δ 15.0, 28.0, 29.8, 43.7, 116.2, 121.6, 122.7, 124.7, 130.4,
130.8, 137.0, 148.2, 162.7; HRESIMS m/z 316.0802 [M + Na]+
(calcd for C15H19NOS2, 316.0800).
N-(2-Acetylphenyl)-3-(ethylthio)propanamide (36). A mixture of
compound 34 (100.0 mg, 0.53 mmol) and sodium ethanethiolate
(54.0 mg, 0.64 mmol) in THF−H2O was heated at 60 °C for 4 h. The
mixture was evaporated under reduced pressure, transferred into a
separatory funnel, and extracted with water (30 mL) and EtOAc (3 ×
(E)-2-(2-Methoxyvinyl)quinolin-4(1H)-one (4) and 3-(Dimethox-
ymethyl)-4-methylquinolin-2(1H)-one (16). Following general pro-
cedure B, with compound 30 (30 mg, 0.12 mmol) as starting material,
G
https://dx.doi.org/10.1021/acs.jnatprod.0c00865
J. Nat. Prod. XXXX, XXX, XXX−XXX

Journal of Natural Products
pubs.acs.org/jnp
Article
yielded compound 4 as a white solid (5 mg, 23%) and compound 16
124.6, 128.5, 130.0, 137.3, 145.4, 163.5; HRESIMS m/z 256.0766 [M
+ Na]+ (calcd for C13H15NOS, 256.0767).
as a pale yellow solid (21 mg, 48%).
1
Compound 4: H NMR (600 MHz, methanol-d4) δ 3.82 (s, 3H),
Dimethyl 2-(Phenylamino)maleate (37). To a solution of aniline
(23) (3.0 g, 32.2 mmol) in MeOH was added dimethylacetylene
dicarboxylate (5.3 g, 37.1 mmol), and the mixture was stirred for 12 h
at reflux. MeOH was evaporated, CH2Cl2 was added (200 mL), and
the mixture was washed with saturated aqueous NH4Cl (2 × 100 mL)
and H2O (2 × 100 mL), dried over MgSO4, and evaporated under
reduced pressure to obtain the product 37 as a yellow oil (7.4 g,
5.79 (d, J = 12.9 Hz, 1H), 6.35 (s, 1H), 7.37 (t, J = 7.7 Hz, 1H), 7.56
(d, J = 8.2 Hz, 1H), 7.66 (d, J = 12.9 Hz, 1H), 7.66 (t, J = 7.7 Hz, 1
H), 8.18 (dd, J = 8.2, 1.1 Hz, 1H); 13C NMR (150 MHz, methanol-
d4) δ 58.2, 99.8, 104.5, 118.9, 124.9, 125.7, 125.9, 133.3, 141.5, 151.4,
157.8, 180.0; HRESIMS m/z 202.0859 [M + H]+ (calcd for
C12H11NO2, 202.0863).
Compound 16: 1H NMR (400 MHz, CDCl3) δ 2.75 (s, 3H), 3.54
(s, 6H), 6.08 (s, 1H), 7.23 (t, J = 7.5 Hz, 1H), 7.41 (t, J = 8.0 Hz,
1H), 7.51 (t, J = 7.5 Hz, 1H), 7.80 (d, J = 8.1, Hz, 1 H), 12.57 (s,
1H); 13C NMR (100 MHz, CDCl3) δ 15.4, 56.2, 102.0, 116.3, 121.5,
122.6, 125.0, 126.7, 130.6, 137.8, 149.1, 163.8; HRESIMS m/z
256.0943 [M + Na]+ (calcd for C13H15NO3, 256.0944).
98%): H NMR analysis revealed the presence of minor impurities
1
remaining; the compound was used without additional purification in
1
subsequent reactions. H NMR (400 MHz, CDCl3) δ 3.64 (s, 3H),
3.69 (s, 3H), 5.38 (s, 1H), 6.87 (d, J = 7.6 Hz, 2H), 7.05 (t, J = 7.4
Hz, 1H), 7.24 (t, J = 7.6 Hz, 2H), 9.68 (s, 1H); 13C NMR data (100
MHz, CDCl3) δ 51.0, 52.5, 93.3, 120.5, 124.0, 129.0, 140.1, 147.8,
164.6, 169.6; HRESIMS m/z 236.0925 [M + H]+ (calcd for
C12H13NO4, 236.0917).
Methyl 4-Oxo-1,4-dihydroquinoline-2-carboxylate (22). A mix-
ture of 37 (5.2 g, 22.1 mmol) and PPA (25.0 g) were heated at 130
°C for 4 h. The resulting slurry was poured into an iced saturated
aqueous solution of NaHCO3. The precipitate was filtered and
washed with H2O and Et2O; the product was obtained as a white solid
(4.1 g, 90%): 1H NMR (400 MHz, DMSO-d6) δ 3.92 (s, 3H), 6.70 (s,
1H), 7.34 (d, J = 7.5 Hz, 1H), 7.65 (t, J = 7.5 Hz, 1H), 7.89 (d, J = 8.4
Hz, 1H), 8.10 (d, J = 8.0 Hz, 1H); 13C NMR data (100 MHz,
DMSO-d6) δ 53.1, 109.3, 121.9, 123.8, 124.5, 126.3, 131.7, 140.5,
142.5, 163.9, 176.1; ESIMS m/z 204.06 [M + H]+.
(E)-2-(2-Ethoxyvinyl)quinolin-4(1H)-one (5) and 3-(Diethoxy-
methyl)- 4-methylquinolin-2(1H)-one (17). Following general
procedure B, with compound 31 (40 mg, 0.17 mmol) as the starting
material, yielded compound 5 as a white solid (10 mg, 28%) and
compound 17 as a pale yellow solid (19 mg, 45%).
Compound 5: 1H NMR (600 MHz, methanol-d4) δ 1.36 (t, J = 7.0
Hz, 3H), 4.04 (q, J = 7.0 Hz, 2H), 5.75 (d, J = 12.8 Hz, 1H), 6.33 (s,
1H), 7.35 (t, J = 7.5 Hz, 1H), 7.54 (d, J = 8.3 Hz, 1H), 7.62 (d, J =
12.8 Hz, 1H), 7.64 (t, J = 7.6 Hz, 1 H), 8.16 (d, J = 8.3 Hz, 1H); 13C
NMR (150 MHz, methanol-d4) δ 15.0, 68.2, 100.2, 104.2, 118.9,
124.9, 125.6, 125.8, 133.2, 141.4, 151.6, 157.2, 179.8; HRESIMS m/z
216.1020 [M + H]+ (calcd for C13H13NO2, 216.1019).
Compound 17: 1H NMR (400 MHz, CDCl3) δ 1.27 (t, J = 7.1 Hz,
6H), 2.78 (s, 3H), 3.62 (dq, J = 9.5, 7.0 Hz, 2H), 3.87 (dq, J = 9.5, 7.1
Hz, 2H), 6.27 (s, 1H), 7.21 (t, J = 7.5 Hz, 1H), 7.40 (d, J = 8.3 Hz,
1H), 7.48 (t, J = 7.5 Hz, 1H), 7.80 (d, J = 8.3 Hz, 1 H), 12.65 (s, 1H);
13C NMR (100 MHz, CDCl3) δ 15.4, 15.5, 63.9, 98.9, 116.3, 121.7,
2-(Hydroxymethyl)quinolin-4(1H)-one (38). To a solution of 22
(1.0 g, 4.9 mmol) in a 1:1 mixture of CH2Cl2−MeOH (20 mL) was
added NaBH4 (0.9 g, 24.5 mmol) at 0 °C. The reaction was left to stir
for 2 h at 0 °C and at rt for an additional 2 h. After this time, EtOAc
(20 mL) was added at 0 °C followed by the addition of MeOH (10
mL), and the mixture was evaporated under reduced pressure. The
residue was purified by silica gel flash chromatography (CH2Cl2−
MeOH, 100:0 to 10:1) to give the product as a white solid (0.4 g,
122.5, 124.9, 127.8, 130.4, 137.8, 148.9, 163.8; HRESIMS m/z
284.1255 [M + Na]+ (calcd for C15H19NO3, 284.1257).
4-Methyl-3-((methylthio)methyl)quinolin-2(1H)-one (12). Fol-
lowing general procedure B, using compound 32 (375 mg, 1.58
mmol) as starting material, compound 12 was obtained as a pale
yellow solid (312 mg, 90%); 1H NMR (400 MHz, CDCl3) δ 2.21 (s,
3H), 2.58 (s, 3H), 3.96 (s, 2H), 7.23 (ddd, J = 8.3, 7.1, 1.2 Hz, 1H),
7.39 (d, J = 8.3 Hz, 1H), 7.49 (ddd, J = 8.3, 7.2, 1.3 Hz, 1H), 7.73
(dd, J = 8.2, 1.0 Hz, 1 H), 11.96 (s, 1H); 13C NMR (100 MHz,
CDCl3) δ 15.5, 16.0, 29.2, 116.3, 121.0, 122.6, 124.7, 128.0, 130.1,
137.2, 145.5, 163.4; HRESIMS m/z 242.0610 [M + Na]+ (calcd for
C12H13NOS, 242.0610).
3-(Methoxymethyl)-4-methylquinolin-2(1H)-one (13). Following
general procedure B, using 33 (535 mg, 2.42 mmol) as a starting
material, compound 13 was obtained as a pale yellow solid (432 mg,
88%): 1H NMR (400 MHz, CDCl3) δ 2.60 (s, 3H), 3.47 (s, 3H), 4.70
(s, 2H), 7.23 (t, J = 7.7 Hz, 1H), 7.35 (d, J = 8.2 Hz, 1H), 7.50 (t, J =
7.7 Hz, 1H), 7.73 (d, J = 8.2 Hz, 1 H), 11.54 (s, 1H); 13C NMR (100
MHz, CDCl3) δ 15.4, 58.5, 65.6, 116.2, 120.9, 122.6, 125.0, 126.7,
130.6, 137.8, 148.8, 163.6; HRESIMS m/z 226.0838 [M + Na]+
(calcd for C12H13NO2, 226.0838).
3-((Dimethylamino)methyl)-4-methylquinolin-2(1H)-one (14).
Following general procedure B, using compound 35 (23 mg, 0.10
mmol) as a starting material, 14 was obtained as a pale yellow solid
(19 mg, 93%): 1H NMR (400 MHz, CDCl3) δ 2.36 (s, 6H), 2.61 (s,
3H), 3.68 (s, 2H), 7.21 (t, J = 7.7 Hz, 1H), 7.40 (d, J = 7.9 Hz, 1H),
7.48 (t, J = 7.7 Hz, 1H), 7.74 (d, J = 8.2 Hz, 1 H), 12.29 (s, 1H); 13C
NMR (100 MHz, CDCl3) δ 15.6, 45.5, 53.8, 116.3, 121.0, 122.4,
124.8, 127.3, 130.1, 137.7, 147.8, 164.3; HRESIMS m/z 217.1336 [M
+ H]+ (calcd for C13H16N2O, 217.1335).
1
48%): H NMR (600 MHz, DMSO-d6) δ 4.50 (d, J = 5.9, 2H), 5.76
(t, J = 5.9, 1H), 6.05 (s, 1H), 7.27 (ddd, J = 8.0, 6.9, 1.1 Hz, 1H), 7.60
(ddd, J = 8.4, 6.9, 1.5 Hz, 1H), 7.67 (d, J = 7.8 Hz, 1H), 8.05 (d, J =
6.9 Hz, 1H), 11.53 (s, 1H); 13C NMR data (150 MHz, DMSO-d6) δ
60.2, 105.4, 118.3, 122.8, 124.8, 125.1, 131.5, 140.0, 153.2, 177.0;
HRESIMS m/z 176.0722 [M + H]+ (calcd for C10H10NO2,
176.0706).
2-(Chloromethyl)quinolin-4(1H)-one (21). To a solution of 38
(0.2 g, 1.1 mmol) in CH2Cl2 was added thionyl chloride (0.83 mL,
11.0 mmol) at 0 °C. The reaction was left to stir at rt for 2 h; then the
mixture was evaporated under reduced pressure. A solution of
saturated NaHCO3 was added to adjust the pH to 10, and the solid
was washed with H2O and dried to obtain the desired product as a
white solid (0.20 g, 98%): 1H NMR (600 MHz, DMSO-d6) δ 4.71 (s,
2H), 6.20 (s, 1H), 7.32 (t, J = 7.4 Hz, 1H), 7.59 (d, J = 8.2 Hz, 1H),
7.66 (ddd, J = 7.6, 6.9, 1.4 Hz, 1H), 8.05 (d, J = 8.7 Hz, 1H), 11.91 (s,
1H); 13C NMR data (150 MHz, DMSO-d6) δ 42.5, 108.9, 118.3,
123.3, 124.8, 132.3, 140.3, 147.7, 155.9, 177.2; ESIMS m/z 194.03
[M + H]+.
Dimethyl 2-(Methyl(phenyl)amino)maleate (40). To a solution of
dimethylacetylene dicarboxylate (6.8 g, 48.3 mmol) in MeOH was
added N-methylaniline (39) (4.5 g, 42.0 mmol) dropwise, and the
mixture was stirred for 15 h at reflux. MeOH was evaporated under
vacuum, CH2Cl2 was added (300 mL), and the mixture was washed
with saturated aqueous NH4Cl (2 × 150 mL) and H2O (2 × 150
mL), dried over MgSO4, and evaporated under reduced pressure. The
residue was purified by silica gel flash chromatography (hexanes−
EtOAc, 10:1 to 5:3) to give compound 40 (9.3 g, 89%) as a yellow
3-((Ethylthio)methyl)-4-methylquinolin-2(1H)-one (15). Follow-
ing general procedure B, using compound 36 (27 mg, 0.11 mmol) as a
starting material, compound 15 was obtained as a pale yellow solid
1
oil: H NMR (400 MHz, CDCl3) δ 3.18 (s, 3H), 3.59 (s, 3H), 3.63
(s, 3H), 4.81 (s, 1H), 7.19 (d, J = 8.1 Hz, 2H), 7.24 (t, J = 7.4 Hz,
1H), 7.34 (t, J = 7.6 Hz, 2H); 13C NMR data (100 MHz, CDCl3) δ
40.3, 50.3, 52.0, 87.7, 126.0, 126.9, 129.0, 144.1, 153.7, 164.7, 167.2;
ESIMS m/z 250.11 [M + H]+.
Methyl 1-Methyl-4-oxo-1,4-dihydroquinoline-2-carboxylate
(41). A mixture of 40 (5.0 g, 20.1 mmol) and PPA (25.0 g) was
1
(23 mg, 92%): H NMR (600 MHz, CDCl3) δ 1.35 (t, J = 7.4 Hz,
3H), 2.57 (s, 3H), 2.70 (q, J = 7.4 Hz, 2H), 4.00 (s, 2H), 7.22 (ddd, J
= 8.3, 7.1, 1.3 Hz, 1H), 7.43 (d, J = 8.3 Hz, 1H), 7.47 (ddd, J = 8.2,
7.1, 1.3 Hz, 1H), 7.71 (dd, J = 8.2, 0.9 Hz, 1 H), 12.63 (s, 1H); 13C
NMR (150 MHz, CDCl3) δ 15.1, 26.8, 27.2, 29.8, 116.4, 121.0, 122.5,
H
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heated at 130 °C for 4 h. The resulting slurry was poured into an iced
saturated aqueous solution of NaHCO3, and the pH was adjusted to 7
by addition of further saturated NaHCO3. The mixture was extracted
with EtOAc (3 × 200 mL), and the combined organics were dried
(MgSO4) and evaporated under reduced pressure. The residue was
purified by silica gel flash chromatography (CH2Cl2−MeOH, 100:0 to
MHz, methanol-d4) δ 14.2, 23.2, 32.5, 33.2, 37.8, 108.9, 119.1, 125.1,
125.3, 125.6, 126.0, 133.4, 136.4, 141.6, 155.6, 180.8; HRESIMS m/z
264.1359 [M + Na]+ (calcd for C16H19NO, 264.1359).
2-Benzyl-1-methylquinolin-4(1H)-one (9). Following general
procedure C, using 43 (14 mg, 0.06 mmol) and 4,4,5,5-tetramethyl-
2-phenyl-1,3,2-dioxaborolane (14 mg, 0.07 mmol), 9 was obtained as
1
1
a white solid (16 mg, 97%): H NMR (600 MHz, CDCl3) δ 3.59 (s,
10:1) to give the product as pink needle crystals (3.2 g, 72%): H
3H), 4.10 (s, 2H), 6.31 (s, 1H), 7.19 (d, J = 7.1 Hz, 2H), 7.26 (t, J =
7.3 Hz, 1H), 7.33 (t, J = 7.4 Hz, 2H), 7.38 (t, J = 7.5 Hz, 1H), 7.44 (t,
J = 8.6 Hz, 1H), 7.64 (ddd, J = 8.7, 7.0, 1.7 Hz, 1H), 8.47 (dd, J = 8.0,
1.6 Hz, 1H); 13C NMR data (100 MHz, CDCl3) δ 34.8, 41.3, 113.6,
115.4, 123.6, 126.9, 126.9, 127.5, 128.4, 129.3, 132.3, 136.1, 142.3,
152.3, 178.1; HRESIMS m/z [M + Na]+ 272.1045 (calcd for
C17H15NO, 272.1046).
1-Methyl-2-(4-(methylthio)benzyl)quinolin-4(1H)-one (10). Fol-
lowing general procedure C, using 43 (14 mg, 0.06 mmol) and
4,4,5,5-tetramethyl-2-((4-methylthio)phenyl)-1,3,2-dioxaborolane
(17 mg, 0.07 mmol), compound 10 was obtained as a white solid (16
mg, 96%): 1H NMR (600 MHz, CDCl3) δ 2.46 (s, 3H), 3.59 (s, 3H),
4.06 (s, 2H), 6.30 (s, 1H), 7.11 (d, J = 8.0 Hz, 2H), 7.21 (d, J = 8.0
Hz, 2H), 7.39 (t, J = 7.4 Hz, 1H), 7.45 (d, J = 8.6 Hz, 1H), 7.65 (t, J =
7.6 Hz, 1H), 8.47 (d, J = 7.8 Hz, 1H); 13C NMR data (150 MHz,
CDCl3) δ 16.0, 34.7, 40.8, 113.5, 115.4, 123.6, 126.8, 126.9, 127.4,
128.8, 132.4, 132.7, 137.8, 142.2, 152.3, 178.1; HRESIMS m/z
318.0923 [M + Na]+ (calcd for C18H17NOS, 318.0923).
NMR (400 MHz, CDCl3) δ 3.76 (s, 3H), 3.96 (s, 3H), 6.57 (s, 1H),
7.34 (t, J = 8.6 Hz, 1H), 7.48 (d, J = 8.5 Hz, 1H), 7.34 (t, J = 5.9 Hz,
1H), 8.33 (d, J = 7.5 Hz, 1H); 13C NMR data (100 MHz, CDCl3) δ
37.3, 53.6, 112.4, 116.1, 124.2, 126.5, 127.1, 133.1, 141.9, 143.8,
164.1, 178.0; HRESIMS m/z 218.0793 [M + H]+ (calcd for
C12H11NO3, 218.0812).
2-(Hydroxymethyl)-1-methylquinolin-4(1H)-one (42). The proce-
dure was the same as for the synthesis of compound 38, using 41 (0.7
g, 3.2 mmol) as starting material. Compound 42 (0.6 g, 74%) was
1
obtained as an orange solid: H NMR (600 MHz, DMSO-d6) δ 3.72
(s, 3H), 4.59 (d, J = 5.8, 2H), 5.76 (t, J = 5.8, 1H), 6.23 (s, 1H), 7.37
(ddd, J = 7.9, 6.8, 1.0 Hz, 1H), 7.72 (ddd, J = 8.6, 6.8, 1.7 Hz, 1H),
7.76 (d, J = 8.4 Hz, 1H), 8.17 (dd, J = 8.0, 1.4 Hz, 1H); 13C NMR
data (150 MHz, DMSO-d6) δ 33.7, 80.9, 108.5, 116.4, 122.9, 125.3,
126.0, 132.1, 141.8, 154.2, 176.3; ESIMS m/z 190.08 [M + H]+.
2-(Chloromethyl)-1-methylquinolin-4(1H)-one (43). The proce-
dure was the same as for the synthesis of compound 21, using 42 (0.2
g, 1 mmol) as starting material. Compound 43 (0.2 g, 90%) was
1
(E)-2-(Hept-2-en-1-yl)-1-methylquinolin-4(1H)-one (11). Follow-
ing general procedure C, using 43 (14 mg, 0.06 mmol) and (E)-hex-1-
en-1-ylboronic acid (9 mg, 0.07 mmol), compound 11 was obtained
obtained as a white solid: H NMR (400 MHz, methanol-d4) δ 3.86
(s, 3H), 4.88 (s, 2H), 6.38 (s 1H), 7.40 (dt, J = 8.0, 4.0 Hz, 1H), 7.73
(d, J = 3.3 Hz, 2H), ;8.21 (d, J = 8.1 Hz, 1H); 13C NMR data (150
MHz, methanol-d4) δ 35.7, 44.2, 112.3, 117.8, 125.4, 126.6, 127.1,
134.4, 143.3, 151.9, 179.9; ESIMS m/z 208.1 [M + H]+.
1
as a white solid (14 mg, 94%): H NMR (600 MHz, CDCl3) δ 0.87
(t, J = 7.1, 3H), 1.25−1.38 (m, 4H), 2.04 (q, J = 6.6 Hz, 2H), 3.41 (d,
J = 4.9 Hz, 2H), 3.71 (s, 3H), 5.46−5.60 (m, 2H), 6.25 (s, 1H), 7.36
(t, J = 7.5 Hz, 1H), 7.49 (d, J = 8.6 Hz, 1H), 7.65 (t, J = 8.4 Hz, 1H),
8.44 (d, J = 7.0 Hz, 1H); 13C NMR data (150 MHz, CDCl3) δ 14.0,
22.3, 31.4, 32.3, 34.5, 38.0, 111.9, 115.4, 123.5, 123.9, 126.7, 126.8,
132.2, 135.4, 142.1, 153.3, 178.1; HRESIMS m/z [M + Na]+
278.1516 (calcd for C17H21NO, 278.1515).
General Procedure C for the Suzuki−Miyaura Cross-
Coupling. A mixture of the chloromethylquinolone (1 equiv),
boronic acid derivative (1.1 equiv), sodium carbonate (4 equiv), and
dichloro(1,1′-bis(diphenylphosphanyl)ferrocene)palladium(II) com-
plex (0.1 equiv) were charged with argon in a sealed flask. 1,4-
Dioxane and H2O (10 mL, 4:1) were added via syringe, and the
reaction was heated at 100 °C for 8−10 h. The mixture was filtrated
through Celite, washed with MeOH, and evaporated under reduced
pressure. The residue was purified by silica gel flash chromatography
(CH2Cl2−MeOH, 100:0 to 10:1).
2-Benzylquinolin-4(1H)-one (6). Following general procedure C,
using the chloromethylquinolone 21 (15 mg, 0.08 mmol) and 4,4,5,5-
tetramethyl-2-phenyl-1,3,2-dioxaboroloane (18 mg, 0.09 mmol),
compound 6 was obtained as a white solid (18 mg, 95%): 1H
NMR (600 MHz, methanol-d4) δ 4.04 (s, 2H), 6.15 (s, 1H), 7.22−
7.33 (m, 5H), 7.36 (ddd, J = 8.1, 7.0, 1.0 Hz, 1H), 7.57 (d, J = 8.0 Hz,
1H), 7.66 (ddd, J = 8.4, 7.0, 1.5 Hz, 1H), 8.19 (d, J = 8.3 Hz, 1H);
13C NMR data (150 MHz, methanol-d4) δ 40.6, 109.8, 119.1, 125.2,
Biological Testing. Procedures for agar well diffusion assays,31
MIC determination and growth inhibition studies,32 time−kill
experiments,31 and swarming motility assays16 were adapted from
existing protocols. Additional details are given in the Supporting
Information.
ASSOCIATED CONTENT
■
sı
* Supporting Information
The Supporting Information is available free of charge at
https://pubs.acs.org/doi/10.1021/acs.jnatprod.0c00865.
1H NMR and 13C spectra for all synthesized compounds,
COSY, HSQC, and HMBC spectra for compound 17,
and full methods for biological assays (PDF)
125.5, 126.0, 128.2, 129.9, 130.0, 133.5, 137.9, 141.6, 155.5, 180.7;
HRESIMS m/z [M + Na]+ 258.0888 (calcd for C16H13NO
258.0889).
2-(4-(Methylthio)benzyl)quinolin-4(1H)-one (7). Following gen-
eral procedure C, using 21 (17 mg, 0.09 mmol) and 4,4,5,5-
tetramethyl-2-((4-methylthio)phenyl)-1,3,2-dioxaborolane (25 mg,
0.10 mmol), compound 7 was obtained as a white solid (24 mg,
98%): 1H NMR (600 MHz, DMSO-d6) δ 2.45 (s, 3H), 3.91 (s, 2H),
5.87 (s, 1H), 7.24 (d, J = 8.4 Hz, 2H), 7.28 (t, J = 7.0 Hz, 2H), 7.30
(d, J = 8.4 Hz, 1H), 7.53 (d, J = 8.2 Hz, 1H), 7.61 (t, J = 8.4 Hz, 1H),
8.02 (d, J = 6.8 Hz, 1H), 11.64 (s, 1H); 13C NMR data (150 MHz,
DMSO-d6) δ 14.7, 38.2, 108.3, 117.8, 122.7, 124.5, 124.6, 126.2,
129.3, 131.4, 133.9, 136.3, 140.0, 152.1, 176.7; HRESIMS m/z
304.0765 [M + Na]+ (calcd for C17H15NOS, 304.0766).
AUTHOR INFORMATION
■
Corresponding Author
Benjamin R. Clark − School of Pharmaceutical Science and
Technology, Tianjin University, Tianjin 300092, People’s
Republic of China; orcid.org/0000-0001-8987-7092;
Phone: +86-22-87401830; Email: bclark@tju.edu.cn
Author
Jianye Li − School of Pharmaceutical Science and Technology,
Tianjin University, Tianjin 300092, People’s Republic of China
(E)-2-(Hept-2-en-1-yl)quinolin-4(1H)-one (8). Following general
procedure C, using 21 (10 mg, 0.05 mmol) and (E)-hex-1-en-1-
ylboronic acid (8 mg, 0.06 mmol), compound 8 was obtained as a
Complete contact information is available at:
https://pubs.acs.org/10.1021/acs.jnatprod.0c00865
1
white solid (12 mg, 93%): H NMR (600 MHz, methanol-d4) δ 0.9
(t, J = 7.2, 3H), 1.31−1.42 (m, 4H), 2.08 (q, J = 7.0 Hz, 2H), 3.41 (d,
J = 6.7 Hz, 2H), 5.56−5.65 (m, 1H), 5.71 (dd, J = 14.4, 7.5 Hz, 1H),
6.21 (s, 1H), 7.38 (t, J = 8.0 Hz, 1H), 7.57 (d, J = 8.3 Hz, 1H), 7.68
(t, J = 7.0 Hz, 1H), 8.21 (d, J = 7.1 Hz, 1H); 13C NMR data (150
Notes
The authors declare no competing financial interest.
I
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(27) Li, D.; Oku, N.; Hasada, A.; Shimizu, M.; Igarashi, Y. Beilstein J.
Org. Chem. 2018, 14, 1446−1451.
ACKNOWLEDGMENTS
■
This work was funded in part by the award from the Research
Fund for International Young Scientists (81850410550) from
the National Natural Science Foundation of China and by a
grant from the National Basic Research Program of China
(2015CB856500). The authors acknowledge Dr. R. Srinivasan,
K. Woycechowsky, and E. Sorensen for their helpful
discussions during the preparation of the manuscript. We
would like to thank Y. Gao and all staff at the analytical center
of the School of Pharmaceutical Science and Technology for
their assistance in the acquisition of MS data.
̈
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