Design, synthesis, and antiviral, fungicidal, and insecticidal activities of tetrahydro-β-carboline-3-carbohydrazide derivatives

Article abstract of DOI:10.1021/jf503794g

According to our previous research on the antiviral activity of β-carboline and tetrahydro-β-carboline derivatives, using (1S,3S)-1-methyl-2,3,4,9-tetrahydro-1H-pyrido[3,4-b]indole-3-carbohydrazide (1) as a lead compound, series of novel tetrahydro-β-carb

Full text of DOI:10.1021/jf503794g

Article
pubs.acs.org/JAFC
Design, Synthesis, and Antiviral, Fungicidal, and Insecticidal
Activities of Tetrahydro-β-carboline-3-carbohydrazide Derivatives
Yongxian Liu,† Hongjian Song,*,† Yuanqiong Huang,† Jiarui Li,‡ Sheng Zhao,§ Yuchuan Song,§
Peiwen Yang,‡ Zhixin Xiao,§ Yuxiu Liu,† Yongqiang Li,† Hui Shang,*,‡ and Qingmin Wang*,†
†State Key Laboratory of Elemento-Organic Chemistry, Research Institute of Elemento-Organic Chemistry, Collaborative Innovation
Center of Chemical Science and Engineering (Tianjin), Nankai University, Tianjin 300071, People’s Republic of China
‡Agricultural Environment and Resources Institute, Yunnan Academy of Agricultural Sciences (YAAS), Yunnan 650205, People’s
Republic of China
§Tobacco Yunnan Company, Yunnan 650011, People’s Republic of China
ABSTRACT: According to our previous research on the antiviral activity of β-carboline and tetrahydro-β-carboline derivatives,
using (1S,3S)-1-methyl-2,3,4,9-tetrahydro-1H-pyrido[3,4-b]indole-3-carbohydrazide (1) as a lead compound, series of novel
tetrahydro-β-carboline derivatives containing acylhydrazone moiety were designed, synthesized, and first evaluated for their
biological activities. Most of these compounds exhibited excellent antiviral activity both in vitro and in vivo. The in vivo
inactivation, curative, and protection activities of compounds 8, 9, 12, 16, 28, 29, and 30 were much higher than that of ribavirin
(37.6%, 39.4%, and 37.9% at 500 μg/mL) and the lead compound (40.0%, 42.3%, and 39.6% at 500 μg/mL). Especially, the in
vitro and in vivo activities of compound 16 (36.9%, 33.6%, 30.2%, and 35.8%) at 100 μg/mL, which were very close to that of
ribavirin (40.0% for in vitro activity) at 500 μg/mL. Compounds 9 and 29 were chosen for the field trials of antiviral efficacy
against TMV (tobacco mosaic virus); the results exhibited that both compounds, especially compound 29, showed better
activities than control plant virus inhibitors. At the same time, the fungicidal results showed that compounds 6, 9, and 11
exhibited good fungicidal activities against 14 kinds of phytopathogens. Additionally, compounds 3 and 23 exhibited moderate
insecticidal activity against the four tested species of insects.
KEYWORDS: tetrahydro-β-carboline, acylhydrazone, antiviral activity, antifungal activity, insecticidal activity, field trials
fragment (−CONHNCH−) widely exist in drug molecules,
INTRODUCTION
■
which show good insecticidal or herbicidal activities such as
Botanical pesticides refer to the pesticides that are developed
using plant resources. They may be the exact active ingredients
extracted from plants or the compounds deried from the active
compounds. Because most of them are safe to untargetted
pymetrozine,14 metaflumizone,15 and diflufenzopyr.16 In this
work, by adopting the tactics of active fragment stitching and
using compound 1 as the lead compound, we designed and
synthesized a series of novel tetrahydro-β-carboline derivatives
beings and they could be rapidly degraded to reduce the
containing acylhydrazone moiety (Figure 2). Meanwhile, their
environment risk of residues, botanical pesticides have long
anti-TMV, fungicidal, and insecticidal activities were evaluated
been touted as attractive alternatives and have become one of
for the first time. Based on the comprehensive analysis of
bioactivity, synthetic procedure, physical properties, and so on,
compounds 9 and 29 were chosen for the field trials of antiviral
efficacy against TMV. Both compounds exhibited better
activities than control plant virus inhibitors, which indicated
that 9 and 29 exhibited a promising application prospect as new
candicates for controlling TMV in the field.
the preferred green biopesticides.1 However, some deficiencies,
such as being too complex to synthesize and active ingredients
being easy to decompose, inevitably limit the widespread use of
botanical pesticides.2,3 Based on the above reasons, our group
was committed to the research on developing novel effective
pesticides using active substances of plant origin as precursors
to solve these problems.
β-Carboline alkaloids are widely distributed in nature. Due to
their broad spectrum of biological activities,4−9 β-carboline
alkaloids have attracted great attention of pharmaceutical
chemists. In our previous work, we found that tetrahydro-β-
carboline-3-carbohydrazide 1 exhibited excellent anti-TMV
activity. The antiviral activity against tobacco mosaic virus
(TMV) in vitro and inactivation, curative, and protection
activities in vivo were 40.4%, 40.0%, 42.3%, and 39.6%,
respectively, at 500 μg/mL.
MATERIALS AND METHODS
■
Instruments. 1H NMR spectra were obtained at 400 MHz using a
Bruker AV400 spectrometer in CDCl3 or DMSO-d6 solution with
tetramethylsilane as the internal standard. HRMS data were obtained
on an FTICR-MS instrument (Ionspec 7.0 T). The melting points
were determined on an X-4 binocular microscope melting point
apparatus and are uncorrected.
Received: August 6, 2014
The biological activities of acylhydrazone compounds had
always been the hot spots on pharmacology research.10−13
Acylhydrazone compounds (Figure 1) containing active
Revised: September 16, 2014
Accepted: September 16, 2014
© XXXX American Chemical Society
A
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Journal of Agricultural and Food Chemistry
Article
Figure 1. Acylhydrazone compounds.
Figure 2. Design of target compounds.
General Synthesis. All anhydrous solvents were dried and purified
by standard techniques. The synthetic routes are given in Scheme 2.
Synthesis of (3S)-1-Methyl-2,3,4,9-tetrahydro-1H-pyrido-
[3,4-b]indole-3-carbohydrazide (1). This compound was synthe-
143.4, 137.9, 135.8, 134.4 and 134.1, 130.0 and 129.8, 128.9 and 128.8,
127.1 and 126.7, 126.9 and 126.9, 120.6, 118.4, 117.5, 111.0, 106.2 and
106.1, 56.7 and 53.3, 48.4 and 47.7, 25.5 and 25.4, 20.0; HRMS (ESI)
calcd for C20H21N4O [M + H]+ 333.1710, found 333.1715.
Compounds 4−26 were synthesized using the similar procedure as
compound 3.
sized by the method17 reported in 99% yield: mp =100−103 °C; H
1
NMR (400 MHz, CDCl3) δ 8.15 (s, 1H, NH), 7.90 (s, 1H, NHCO),
3
3
7.50 (d, JHH = 7.6 Hz, 1H, Ar−H), 7.32 (d, JHH = 8.0 Hz, 1H, Ar−
Data for 1-Methyl-9H-b-carboline-3-carboxylic Acid Benzylidene-
hydrazide (4). This compound was obtained as a yellow solid in 91%
yield: mp = 303−306 °C; 1H NMR (300 MHz, DMSO-d6) δ 12.05 (s,
1H, NH), 11.85 (s, 1H, OC-NH), 8.82 (s, 1H, NCH), 8.69 (s,
3
3
H), 7.17 (t, JHH = 7.2 Hz, 1H, Ar−H), 7.11 (t, JHH = 7.2 Hz, 1H,
Ar−H), 4.18−4.23 (m, 1H, CHCH3), 3.93 (br, 2H, NH2), 3.65 (q,
3JHH = 8.4 Hz, 1H, CHCO), 3.26−3.32 (m, 1H, CH2), 2.71−2.78 (m,
3
3
1H, CH2), 1.48 (d, JHH = 6.8 Hz, 3H, CH3); 13C NMR (100 MHz,
3
1H, Ar−H), 8.41 (d, JHH = 7.8 Hz, 1H, Ar−H), 7.77 (d, JHH = 6.3
Hz, 2H, Ph-H), 7.67 (d, 3JHH = 8.1 Hz, 1H, Ar−H), 7.60 (t, 3JHH = 7.2
Hz, 1H, Ar−H), 7.55−7.41 (m, 3H, Ph-H), 7.31 (t, 3JHH = 7.2 Hz, 1H,
Ar−H), 2.92 (s, 3H, CH3); 13C NMR (100 MHz, DMSO-d6) δ 161.2,
147.9, 141.1, 140.8, 138.2, 136.1, 134.6, 129.9, 128.9, 128.4, 127.5,
127.1, 122.3, 121.4, 120.1, 113.2, 112.3, 20.4; HRMS (ESI) calcd for
C20H17N4O [M + H]+ 329.1397, found 329.1402.
DMSO-d6) δ 172.0, 137.9, 135.8, 126.9, 120.5, 118.3, 117.4, 110.9,
106.2, 56.1, 48.2, 25.4, 20.0; HRMS (ESI) calcd for C13H17N4O [M +
H]+ 245.1397, found 245.1398.
Synthesis of 1-Methyl-9H-carbazole-3-carboxylic Acid Hy-
drazide (2). A mixture of ethyl 1-methyl-9H-pyrido[3,4-b]indole-3-
carboxylate18 (2.00 g, 7.86 mmol) and hydrazine hydrate (80%) (3.15
g, 5.04 mmol) in a mixed solvent of alcohol (80 mL) and DMF (25
mL) was heated at reflux for 24 h; then the mixture was concentrated
in vacuo. Solid was precipitated out and collected by filtration. The
cake was extracted with boiling alcohol (each 30 mL) for several times
until the filter cake did not dissolve in hot alcohol any more. The
combined extract was concentrated to give 2 as a light-pink solid (0.90
g, 48%): mp =242−244 °C; 1H NMR (300 MHz, DMSO-d6) δ 11.92
(s, 1H, NH), 9.51 (s, 1H, OC-NH), 8.64 (s, 1H, Ar−H), 8.35 (d,
3JHH = 8.1 Hz, 1H, Ar−H)), 7.64 (d, 3JHH = 8.1 Hz, 1H, Ar−H), 7.57
Data for N′-Benzylidene-(3R)-1-methyl-2,3,4,9-tetrahydro-1H-
pyrido[3,4-b]indole-3-carbohydrazide (5). This compound was
obtained as a light yellow solid in 76% yield: mp = 197−200 °C;
1H NMR (400 MHz, DMSO-d6) δ 11.67 and 11.47 (s, 1H, NH),
10.98 and 10.86 (s, 1H, OC−NH), 8.35 and 8.09 (s, 1H, NCH),
7.75−7.69 (m, 1H, Ph-H), 7.66−7.60 (m, 1H, Ph-H), 7.50−7.41 (m,
2H, Ph-H), 7.41−7.36 (m, 2H, Ph-H and Ar−H), 7.33 and 7.31 (d,
3JHH = 8.8 Hz, 1H, Ph-H), 7.06 and 7.04 (t, 3JHH = 7.2 Hz, 1H, Ar−H),
3
7.00−6.92 (m, 1H, Ar−H), 4.70−4.61 and 3.69 (dd, JHH = 10.4 Hz,
(t, 3JHH = 7.2 Hz, 1H, Ar−H), 7.28 (t, 3JHH = 7.2 Hz, 1H, Ar−H), 4.55
(s, 2H, NH2), 2.82 (s, 3H, CH3); 13C NMR (100 MHz, DMSO-d6) δ
164.1, 141.0, 140.7, 138.7, 135.7, 128.3, 127.3, 122.2, 121.4, 119.9,
112.2, 111.9, 20.4; HRMS (ESI) calcd for C13H13N4O [M + H]+
241.1084, found 241.1084.
2
3.6 Hz, 1H, CH), 4.42−4.14 (m, 1H, CH), 3.08 and 2.98 (dd, JHH
=
14.8 Hz, 3JHH = 2.8 Hz, 1H, CH), 2.79−2.67 (m, 1H, CH2), 1.53 and
1.48 (d, 3JHH = 6.8 Hz, 3H, CH3); 13C NMR (100 MHz, DMSO-d6) δ
173.4 and 168.8, 147.2 and 144.1, 137.2, 136.0 and 135.9, 134.3 and
134.0, 130.0 and 130.0, 128.8, 127.1, 126.8 and 126.6, 120.9 and 120.7,
118.6 and 118.5, 117.7 and 117.5, 111.1 and 111.1, 106.0 and 105.7,
56.6 and 53.6, 48.5 and 48.1, 25.3 and 24.7, 19.7 and 19.0; HRMS
(ESI) calcd for C20H21N4O [M + H]+ 333.1710, found 333.1708.
Data for N′-(4-tert-Butylbenzylidene)-(3S)-1-methyl-2,3,4,9-tetra-
hydro-1H-pyrido[3,4-b]indole-3-carbohydrazide (6). This com-
pound was obtained as a yellow solid in 72% yield: mp = 139−143
°C; 1H NMR (400 MHz, DMSO-d6) δ 11.47 and 11.32 (s, 1H, NH),
10.87 and 10.82 (s, 1H, OC-NH), 8.31 and 8.03 (s, 1H, NCH),
7.88 and 7.70−7.27 (m, 6H, Ph-H and Ar−H), 7.04 (t, 3JHH = 6.8 Hz,
Synthesis of N′-Benzylidene-(3S)-1-methyl-2,3,4,9-tetrahy-
dro-1H-pyrido[3,4-b]indole-3-carbohydrazide (3). To a slurry of
compound 1 (0.50 g, 2.05 mmol) in toluene (35 mL) was added
dropwise benzaldehyde (0.44 g, 4.10 mmol) at room temperature,
Then the reaction mixture was heated to reflux. After 5 h, TLC
analysis showed the complete consumption of compound 1. The
reaction solution was cooled to room temperature, and solid was
precipitated out. The solid was filtrated and washed with toluene and
then stirred in absolute ether for 1.5 h to remove the remaining
toluene. Filtered once again to give 3 as a yellow solid (0.50 g, 74%):
3
1H, Ar−H), 6.96 (t, JHH = 7.2 Hz, 1H, Ar−H), 4.48 and 3.62 (dd,
1
3JHH = 10.4 Hz, 7.2 Hz, 1H, CH), 4.27−4.08 (m, 1H, CH), 3.02−2.88
mp = 200−204 °C; H NMR (400 MHz, DMSO-d6) δ 11.48 and
11.35 (s, 1H, NH), 10.84 and 10.81 (s, 1H, OC-NH), 8.34 and 8.06
(s, 1H, NCH), 7.71 and 7.61 (dd, 3JHH = 7.6 Hz, 4JHH = 1.6 Hz, 1H,
Ph-H), 7.49−7.42 (m, 2H, Ph-H), 7.42−7.33 (m, 2H, Ph-H and Ar−
(m, 1H, CH2), 2.77−2.59 (m, 1H, CH2), 1.52−1.35 (m, 3H, CH3),
1.30 and 1.24 (s, 9H, CH3); 13C NMR (100 MHz, DMSO-d6) δ 174.3
and 168.9, 152.8 and 152.6, 147.0 and 143.3, 137.8, 135.8, 131.6 and
131.4, 129.2, 126.9 and 126.5, 125.7 and 125.6, 120.6 and 120.6, 118.4,
117.5 and 117.5, 111.0, 106.2 and 106.0, 56.7 and 53.5, 48.4 and 47.7,
34.6 and 34.5, 31.0 and 30.9, 25.5 and 25.2, 19.9 and 19.9; HRMS
(ESI) calcd for C24H29N4O [M + H]+ 389.2336, found 389.2338.
Data for N′-(4-(Dimethylamino)benzylidene)-(3S)-1-methyl-
2,3,4,9-tetrahydro-1H-pyrido[3,4-b]indole-3-carbohydrazide (7).
3
3
H), 7.31 and 7.30 (d, JHH = 8.0 Hz, 1H, Ar−H), 7.03 (t, JHH = 7.2
Hz, 1H, Ar−H), 6.95 and 6.92 (t, 3JHH = 7.2 Hz, 1H, Ar−H), 4.48 and
3.61 (dd, 3JHH = 10.8 Hz, 3.6 Hz, 1H, CH), 4.15 (q, 3JHH = 8.0 Hz, 1H,
2
CH), 2.98−2.89 (m, 1H, CH2), 2.71 and 2.62 (ddd, JHH = 14.4 Hz,
3JHH = 11.6 Hz, 4JHH = 2.0 Hz, 1H, CH2), 1.45 (d, 3JHH = 6.8 Hz, 3H,
CH3); 13C NMR (100 MHz, DMSO-d6) δ 174.6 and 169.1, 147.0 and
B
dx.doi.org/10.1021/jf503794g | J. Agric. Food Chem. XXXX, XXX, XXX−XXX

Journal of Agricultural and Food Chemistry
Article
3
3
This compound was obtained as a yellow solid in 73% yield: mp =
215−220 °C; H NMR (400 MHz, DMSO-d6) δ 11.20 and 11.05 (s,
and 7.39 (d, JHH = 8.4 Hz, 1H, Ar−H), 7.33 and 7.31 (d, JHH = 8.4
Hz, 1H, Ar−H), 7.06 and 7.04 (t, 3JHH = 7.2 Hz, 1H, Ar−H), 6.96 and
6.96 (t, 3JHH = 7.2 Hz, 1H, Ar−H), 4.68 and 3.70 (dd, 3JHH = 10.4 Hz,
4.4 Hz, 1H, CH), 4.38 and 4.19 (q, 3JHH = 6.4 Hz, 1H, CH), 3.06 and
2.98 (dd, 2JHH = 14.4 Hz, 3JHH = 3.6 Hz, 1H, CH2), 2.80−2.66 (m, 1H,
1
1H, NH), 10.84 and 10.80 (s, 1H, OC-NH), 8.16 and 7.92 (s, 1H,
NCH), 7.52 and 7.36 (d, 3JHH = 7.6 Hz, 2H, Ph-H), 7.44−7.38 (m,
1H, Ar−H), 7.30 (d, 3JHH = 6.4 Hz, 1H, Ar−H), 7.03 (t, 3JHH = 6.8 Hz,
1H, Ar−H), 6.95 (t, 3JHH = 6.8 Hz, 1H, Ar−H), 6.75 and 6.67 (d, 3JHH
= 7.6 Hz, 2H, Ph-H), 4.44 and 3.57 (d, 3JHH = 8.8 Hz, 1H, CH), 4.23−
4.08 (m, 1H, CH), 3.07−2.86 (m, 7H, N−CH3 and CH2), 2.74−2.56
(m, 1H, CH2), 1.52−1.38 (m, 3H, CH3); 13C NMR (100 MHz,
DMSO-d6) δ 173.8 and 168.5, 151.5 and 151.3, 147.8 and 144.2,
137.8, 135.8, 128.4 and 127.9, 126.9, 121.6 and 121.5, 120.6 and 120.5,
118.4, 117.5, 111.9 and 111.8, 111.0, 106.2 and 106.1, 56.6 and 53.5,
48.4 and 47.7, 39.8, 25.6 and 25.2, 19.9; HRMS (ESI) calcd for
C22H26N5O [M + H]+ 376.2132, found 376.2137.
CH2), 1.53 and 1.48 (d, JHH = 6.4 Hz, 3H, CH3); 13C NMR (100
3
MHz, DMSO-d6) δ 173.5 and 169.0, 144.5 and 141.6, 137.2, 135.9 and
135.9, 135.2 and 134.8, 132.2 and 132.1, 131.7, 131.1, 128.6 and 128.5,
126.9 and 126.8, 126.6 and 126.4, 120.9 and 120.7, 118.6 and 118.5,
117.7 and 117.5, 111.1 and 111.0, 105.9 and 105.6, 56.6 and 53.5, 48.5
and 48.1, 25.3 and 24.7, 19.6 and 18.9; HRMS (ESI) calcd for
C20H19N4OCl2 [M + H]+ 401.0931, found 401.0934.
Data for N′-(4-Methoxybenzylidene)-(3S)-1-methyl-2,3,4,9-tetra-
hydro-1H-pyrido[3,4-b]indole-3-carbohydrazide (12). This com-
pound was obtained as a yellow solid in 69% yield: mp = 138−143
°C; 1H NMR (400 MHz, DMSO-d6) δ 11.35 and 11.22 (s, 1H, NH),
10.84 and 10.81 (s, 1H, OC-NH), 8.27 and 8.00 (s, 1H, NCH),
Data for N′-(4-Nitrobenzylidene)-(3S)-1-methyl-2,3,4,9-tetrahy-
dro-1H-pyrido[3,4-b]indole-3-carbohydrazide (8). This compound
1
was obtained as a yellow solid in 74% yield: mp = 222−227 °C; H
3
3
NMR (400 MHz, DMSO-d6) δ 11.81 and 11.68 (s, 1H, NH), 10.87
and 10.82 (s, 1H, OC-NH), 8.46 and 8.17 (s, 1H, NCH), 8.31
7.65 and 7.55 (d, JHH = 8.0 Hz, 2H, Ph-H), 7.40 and 7.36 (d, JHH
=
3
7.6 Hz, 1H, Ar−H), 7.30 (d, JHH = 7.6 Hz, 1H, Ar−H), 7.08−6.89
(m, 4H, Ar−H and Ph-H), 4.45 and 3.59 (d, 3JHH = 8.8 Hz, 1H, CH),
4.20−4.08 (m, 1H, CH), 3.81 and 3.74 (s, 3H, O−CH3), 2.93 (d, 2JHH
3
3
and 8.22 (d, JHH = 8.4 Hz, 2H, Ph-H), 7.98 and 7.88 (d, JHH = 8.4
Hz, 2H, Ph-H), 7.41 and 7.36 (d, 3JHH = 7.6 Hz, 1H, Ar−H), 7.31 (d,
3JHH = 6.4 Hz, 1H, Ar−H), 7.04 (t, 3JHH = 7.2 Hz, 1H, Ar−H), 6.96 (t,
3JHH = 7.6 Hz, 1H, Ar−H), 4.55 and 3.67 (d, 3JHH = 8.8 Hz, 1H, CH),
3
= 14.4 Hz, 1H, CH2), 2.76−2.56 (m, 1H, CH2), 1.45 (d, JHH = 6.0
Hz, 3H, CH3); 13C NMR (100 MHz, DMSO-d6) δ 174.3 and 168.8,
160.8 and 160.6, 146.9 and 143.2, 137.9, 135.8 and 135.8, 128.6 and
128.2, 127.0 and 126.9, 126.7, 120.5, 118.4, 117.5, 114.3, 111.0, 106.2
and 106.1, 56.7 and 53.3, 55.3 and 55.2, 48.4 and 47.6, 25.5 and 25.4,
20.0 and 19.9; HRMS (ESI) calcd for C21H23N4O2 [M + H]+
363.1816, found 363.1819.
2
4.25−4.10 (m, 1H, CH), 2.96 (d, JHH = 13.6 Hz, 1H, CH2), 2.79−
2.60 (m, 1H, CH2), 1.47 (d, 3JHH = 6.4 Hz, 3H, CH3); 13C NMR (100
MHz, DMSO-d6) δ 174.8 and 169.6, 147.8 and 147.7, 144.6, 140.7 and
140.4, 137.8 and 137.5, 135.9, 128.0 and 127.7, 126.9, 124.1, 120.7 and
120.6, 118.5, 117.6 and 117.5, 111.0, 106.1 and 106.0, 56.8 and 53.2,
48.5 and 47.8, 25.4 and 25.4, 19.9 and 19.8; HRMS (ESI) calcd for
C20H20N5O3 [M + H]+ 378.1561, found 378.1563.
Data for N′-(3-Methoxybenzylidene)-(3S)-1-methyl-2,3,4,9-tetra-
hydro-1H-pyrido[3,4-b]indole-3-carbohydrazide (13). This com-
pound was obtained as yellow solid in 63% yield: mp = 186−190
°C; 1H NMR (400 MHz, DMSO-d6) δ 11.54 and 11.39 (s, 1H, NH),
10.87 and 10.83 (s, 1H, OC-NH), 8.32 and 8.03 (s, 1H, NCH),
7.45−7.11 (m, 5H, Ph-H and Ar−H), 7.08−6.89 (m, 3H, Ar−H and
Ph-H), 4.48 and 3.63 (d, 3JHH = 8.0 Hz, 1H, CH), 4.25−4.10 (m, 1H,
CH), 3.81 and 3.69 (s, 3H, O−CH3), 3.02−2.89 (m, 1H, CH2), 2.77−
2.60 (m, 1H, CH2), 1.46 (d, 3JHH = 5.2 Hz, 3H, CH3); 13C NMR (100
MHz, DMSO-d6) δ 174.3 and 169.2, 159.6, 147.1 and 143.4, 137.8 and
137.5, 135.9 and 135.8, 135.6, 130.0 and 130.0, 126.9, 120.7 and 120.6,
120.0 and 119.1, 118.4, 117.5, 116.2 and 115.4, 112.0 and 111.3, 111.0,
106.2 and 106.0, 56.7 and 53.6, 55.2 and 55.1, 48.5 and 47.8, 25.5 and
25.2, 19.9 and 19.8; HRMS (ESI) calcd for C21H23N4O2 [M + H]+
363.1816, found 363.1818.
Data for N′-(4-Chlorobenzylidene)-(3S)-1-methyl-2,3,4,9-tetrahy-
dro-1H-pyrido[3,4-b]indole-3-carbohydrazide (9). This compound
1
was obtained as a yellow solid in 81% yield: mp = 140−145 °C; H
NMR (400 MHz, DMSO-d6) δ 11.62 and 11.51 (s, 1H, NH), 10.91
and 10.85 (s, 1H, OC-NH), 8.35 and 8.07 (s, 1H, NCH), 7.74
and 7.64 (d, 3JHH = 7.2 Hz, 2H, Ph-H), 7.58−7.35 (m, 3H, Ph-H and
Ar−H), 7.35−7.28 (m, 1H, Ar−H), 7.10−7.00 (m, 1H, Ar−H), 7.00−
6.90 (m, 1H, Ar−H), 4.56 and 3.66 (d, 3JHH = 9.6 Hz, 1H, CH), 4.30−
4.12 (m, 1H, CH), 3.03−2.90 (m, 1H, CH2), 2.79−2.62 (m, 1H,
CH2), 1.54−1.39 (m, 3H, CH3); 13C NMR (100 MHz, DMSO-d6) δ
174.2 and 169.1, 145.8 and 142.4, 137.6 and 135.9, 134.5 and 134.3,
133.3 and 133.0, 129.0, 128.7, 128.4, 126.9 and 126.8, 120.7 and 120.6,
118.5 and 118.4, 117.6 and 117.5, 111.0, 106.1 and 105.9, 56.7 and
53.3, 48.5 and 47.8, 25.4 and 25.1, 19.8 and 19.6; HRMS (ESI) calcd
for C20H20N4OCl [M + H]+ 367.1320, found 367.1323.
Data for N′-(2-Methoxybenzylidene)-(3S)-1-methyl-2,3,4,9-tetra-
hydro-1H-pyrido[3,4-b]indole-3-carbohydrazide (14). This com-
pound was obtained as yellow solid in 82% yield: mp = 180−183
°C; 1H NMR (400 MHz, DMSO-d6) δ 11.52 and 11.46 (s, 1H, NH),
10.90 and 10.84 (s, 1H, OC-NH), 8.66 and 8.41 (s, 1H, NCH),
Data for N′-(2,4-Dichlorobenzylidene)-(3S)-1-methyl-2,3,4,9-tet-
rahydro-1H-pyrido[3,4-b]indole-3-carbohydrazide (10). This com-
pound was obtained as a light yellow solid in 85% yield: mp = 211−
1
3
3
213 °C; H NMR (400 MHz, DMSO-d6) δ 11.82 and 11.77 (s, 1H,
7.84 and 7.68 (d, JHH3 = 7.2 Hz, 1H, Ph-H), 7.41 (d, JHH = 7.6 Hz,
3
NH), 10.91 and 10.84 (s, 1H, OC-NH), 8.71 and 8.42 (s, 1H, N
1H, Ar−H), 7.37 (d, JHH = 7.6 Hz, 1H, Ar−H), 7.31 (dd, JHH = 8.0
3
CH), 8.00 and 7.85 (d, JHH = 8.4 Hz, 1H, Ph-H), 7.73 and 7.71 (d,
Hz, 4.0 Hz, 1H, Ph-H), 7.14−6.86 (m, 4H, Ph-H and Ar−H), 4.54 and
4JHH = 2.0 Hz, 1H, Ph-H), 7.53 and 7.43−7.35 (dd, 3JHH = 8.4 Hz, 4JHH
3.61 (dd, JHH = 10.0 Hz, 2.8 Hz, 1H, CH), 4.30−4.10 (m, 1H, CH),
3
3
3.86 and 3.84 (s, 3H, O−CH3), 3.04−2.89 (m, 1H, CH2), 2.77−2.60
(m, 1H, CH2), 1.55−1.37 (m, 3H, CH3); 13C NMR (100 MHz,
DMSO-d6) δ 173.9 and 168.9, 157.7 and 157.6, 142.5 and 139.3,
137.6, 135.8, 131.5 and 131.4, 126.9 and 126.8, 125.6 and 125.2, 122.3
and 122.1, 120.7 and 120.7, 120.6, 118.5 and 118.4, 117.6 and 117.5,
111.8, 111.0, 106.1 and 105.9, 56.6 and 53.5, 55.7, 48.4 and 47.8, 25.5
and 25.1, 19.9 and 19.6; HRMS (ESI) calcd for C21H23N4O2 [M + H]+
363.1816, found 363.1823.
= 2.0 Hz, 1H, Ph-H), 7.41 and 7.37 (d, JHH = 8.0 Hz, 1H, Ar−H),
3
3
7.32 and 7.31 (d, JHH = 8.0 Hz, 1H, Ar−H), 7.04 (t, JHH = 7.2 Hz,
3
1H, Ar−H), 6.96 and 6.94 (t, JHH = 7.2 Hz, 1H, Ar−H), 4.65−4.56
and 3.66 (dd, JHH = 10.4 Hz, 3.6 Hz, 1H, CH), 4.35−4.13 (m, 1H,
3
CH), 3.06−2.91 (m, 1H, CH2), 2.78−2.64 (m, 1H, CH2), 1.54−1.43
(m, 3H, CH3); 13C NMR (100 MHz, DMSO-d6) δ 174.0 and 169.2,
142.0 and 139.0, 137.5 and 137.5, 135.9, 135.0 and 134.9, 133.8 and
133.7, 130.8 and 130.4, 129.4 and 129.4, 128.1 and 128.1, 128.1 and
127.8, 126.8 and 126.7, 120.8 and 120.6, 118.5 and 118.5, 117.7 and
117.5, 111.1 and 111.0, 106.1 and 105.8, 56.7 and 53.4, 48.5 and 48.0,
25.3 and 25.0, 19.8 and 19.3; HRMS (ESI) calcd for C20H19N4OCl2
[M + H]+ 401.0931, found 401.0929.
Data for N′-(3,4-Dimethoxybenzylidene)-(3S)-1-methyl-2,3,4,9-
tetrahydro-1H-pyrido[3,4-b]indole-3-carbohydrazide (15). This
compound was obtained as light yellow solid in 91% yield: mp =
1
203−206 °C; H NMR (400 MHz, DMSO-d6) δ 11.47 and 11.27 (s,
1H, NH), 10.89 and 10.83 (s, 1H, OC-NH), 8.25 and 7.99 (s, 1H,
NCH), 7.43−7.36 (m, 1H, Ar−H), 7.34−7.27 and 7.22−7.15 (m,
3H, Ar−H and Ph-H), 7.07−6.91 (m, 3H, Ar−H and Ph-H), 4.49 and
Data for N′-(3,4-Dichlorobenzylidene)-(3S)-1-methyl-2,3,4,9-tet-
rahydro-1H-pyrido[3,4-b]indole-3-carbohydrazide (11). This com-
pound was obtained as a light yellow solid in 79% yield: mp = 189−
1
3
193 °C; H NMR (400 MHz, DMSO-d6) δ 11.84 and 11.67 (s, 1H,
3.67−3.59 (dd, JHH = 11.2 Hz, 3.6 Hz, 1H, CH), 4.33−4.11 (m, 1H,
NH), 10.98 and 10.86 (s, 1H, OC-NH), 8.34 and 8.07 (s, 1H, N
CH), 3.82 and 3.75 (s, 3H, O−CH3), 3.81 and 3.64 (s, 3H, O−CH3),
2
3
CH), 7.94 and 7.88 (s, 1H, Ph-H), 7.72 and 7.64 (s, 2H, Ph-H), 7.41
3.03 and 2.94 (dd, JHH = 14.8 Hz, JHH = 2.8 Hz, 1H, CH2), 2.76−
C
dx.doi.org/10.1021/jf503794g | J. Agric. Food Chem. XXXX, XXX, XXX−XXX

Journal of Agricultural and Food Chemistry
Article
3
3
2.65 (m, 1H, CH2), 1.48 and 1.46 (d, JHH = 6.8 Hz, 3H, CH3); 13C
2H, Py-H), 8.35 and 8.05 (s, 1H, NCH), 7.65 and 7.57 (d, JHH
=
NMR (100 MHz, DMSO-d6) δ 173.7 and 168.8, 150.7 and 150.5,
149.1 and 148.9, 147.4 and 143.7, 137.7, 135.9, 127.0, 126.9 and 126.9,
121.8 and 120.7, 120.6 and 120.4, 118.4, 117.5, 111.7 and 111.5, 111.0,
109.2 and 108.3, 106.1 and 106.0, 56.6 and 53.8, 55.6 and 55.5, 55.5
and 55.3, 48.4 and 48.0, 25.5 and 24.9, 19.9 and 19.6; HRMS (ESI)
calcd for C22H25N4O3 [M + H]+ 393.1921, found 393.1918.
5.6 Hz, 2H, Py-H), 7.41 and 7.37 (d, 3JHH = 7.6 Hz, 1H, Ph-H), 7.34−
7.29 (m, 1H, Ar−H), 7.04 (t, 3JHH = 7.6 Hz, 1H, Ar−H), 6.95 (t, 3JHH
= 7.6 Hz, 1H, Ar−H), 4.58 and 3.67 (dd, 3JHH = 10.8 Hz, 3.6 Hz, 1H,
3
CH), 4.26 and 4.16 (q, JHH = 6.4 Hz, 1H, CH), 3.05−2.91 (m, 1H,
3
CH2), 2.80−2.62 (m, 1H, CH2), 1.48 (d, JHH = 6.4 Hz, 3H, CH3);
13C NMR (100 MHz, DMSO-d6) δ 174.5 and 169.4, 150.3, 144.7,
Data for N′-(Benzo[d][1,3]dioxol-5-ylmethylene)-(3S)-1-methyl-
2,3,4,9-tetrahydro-1H-pyrido[3,4-b]indole-3-carbohydrazide (16).
This compound was obtained as yellow solid in 83% yield: mp =
199−203 °C; H NMR (400 MHz, DMSO-d6) δ 11.41 and 11.26 (s,
141.6, 141.3 and 141.2, 137.7 and 135.8, 126.9 and 126.8, 121.0, 120.7
and 120.6, 118.5 and 118.4, 117.6 and 117.5, 111.0, 106.0 and 105.9,
56.7 and 53.3, 48.5 and 47.9, 25.4 and 25.2, 19.8 and 19.6; HRMS
(ESI) calcd for C19H20N5O [M + H]+ 334.1662, found 334.1663.
Data for N′-(pyridin-3-ylmethylene)-(3S)-1-methyl-2,3,4,9-tetra-
hydro-1H-pyrido[3,4-b]indole-3-carbohydrazide (20). This com-
pound was obtained as yellow solid in 72% yield: mp = 205−209
°C; 1H NMR (400 MHz, DMSO-d6) δ 11.74 and 11.58 (s, 1H, NH),
10.91 and 10.84 (s, 1H, OC-NH), 8.84 and 8.79 (s, 1H, Py-H), 8.62
and 8.55 (d, 3JHH = 4.0 Hz, 1H, Py-H), 8.41 and 8.12 (s, 1H, NCH),
1
1H, NH), 10.86 and 10.81 (s, 1H, OC-NH), 8.24 and 7.96 (s, 1H,
NCH), 7.40 and 7.36 (d, 3JHH = 7.6 Hz, 1H, Ar−H), 7.31 and 7.30
(d, 3JHH = 8.0 Hz, 1H, Ar−H), 7.28 and 7.17 (s, 1H, Ph-H), 7.15 and
7.09 (d, 3JHH = 8.0 Hz, 1H, Ph-H), 7.06−6.96 (m, 2H, Ar−H and Ph-
H), 6.94 (t, 3JHH = 7.6 Hz, 1H, Ar−H), 6.10 (s, 1H, O−CH2), 6.02 (d,
3
2JHH = 4.8 Hz, 1H, O−CH2), 4.49 and 3.60 (dd, JHH = 10.8 Hz, 3.6
3
8.12 and 8.03 (d, JHH = 4.0 Hz, 1H, Py-H), 7.49 and 7.44−7.35 (m,
Hz, 1H, CH), 4.22 and 4.13 (m, 1H, CH), 2.99−2.87 (m, 1H, CH2),
2.75−2.57 (m, 1H, CH2), 1.45 (d, 3JHH = 6.4 Hz, 3H, CH3); 13C NMR
(100 MHz, DMSO-d6) δ 174.2 and 168.9, 149.1 and 148.9, 148.0 and
147.9, 146.9 and 143.2, 137.8, 135.8, 128.8 and 128.6, 126.9, 123.3 and
122.8, 120.6 and 120.6, 118.4, 117.5, 111.0, 108.5, 106.2 and 106.0,
105.1 and 105.0, 101.6 and 101.5, 56.7 and 53.3, 48.4 and 47.7, 25.5
and 25.2, 19.9 and 19.8; HRMS (ESI) calcd for C21H21N4O3 [M + H]+
377.1608, found 377.1615.
3
2H, Py-H and Ar−H), 7.33 and 7.31 (d, JHH = 7.6 Hz, 1H, Ph-H),
7.04 (t, 3JHH = 7.6 Hz, 1H, Ar−H), 6.96 (t, 3JHH = 7.6 Hz, 1H, Ar−H),
4.59 and 3.66 (dd, 3JHH = 10.8 Hz, 3.6 Hz, 1H, CH), 4.32 and 4.12 (m,
1H, CH), 3.05−2.92 (m, 1H, CH2), 2.78−2.63 (m, 1H, CH2), 1.53−
1.39 (m, 3H, CH3); 13C NMR (100 MHz, DMSO-d6) δ 169.1, 150.7
and 150.5, 148.7 and 148.5, 144.4 and 140.9, 137.6 and 135.8, 133.4
and 133.2, 130.3, 130.0, 126.9 and 126.8, 124.0 and 124.0, 120.7 and
120.6, 118.5 and 118.4, 117.6 and 117.5, 111.0, 106.0 and 105.9, 56.7
and 53.4, 48.5 and 47.9, 25.4 and 25.1, 19.8 and 19.5; HRMS (ESI)
calcd for C19H20N5O [M + H]+ 334.1662, found 334.1664.
Data for N′-((2,3-Dihydrobenzo[b][1,4]dioxin-6-yl)methylene)-
(3S)-1-methyl-2,3,4,9-tetrahydro-1H-pyrido[3,4-b]indole-3-carbo-
hydrazide (17). This compound was obtained as yellow solid in 81%
1
Data for N′-(Pyridin-2-ylmethylene)-(3S)-1-methyl-2,3,4,9-tetra-
hydro-1H-pyrido[3,4-b]indole-3-carbohydrazide (21). This com-
pound was obtained as yellow solid in 85% yield: mp = 245−249
°C; 1H NMR (400 MHz, DMSO-d6) δ 11.69 and 11.65 (s, 1H, NH),
yield: mp = 204−207 °C; H NMR (400 MHz, DMSO-d6) δ 11.44
and 11.26 (s, 1H, NH), 10.89 and 10.82 (s, 1H, OC-NH), 8.21 and
3
7.94 (s, 1H, NCH), 7.40 and 7.36 (d, JHH = 7.6 Hz, 1H, Ar−H),
7.32 and 7.30 (d, 3JHH = 8.0 Hz, 1H, Ar−H), 7.20 and 7.10 (s, 1H, Ph-
10.85 and 10.82 (s, 1H, OC-NH), 8.62 and 8.57 (d, 3JHH = 4.8 Hz,
3
H), 7.19 and 7.09 (d, JHH = 7.6 Hz, 1H, Ph-H), 7.07−7.01 (m, 1H,
3
Ar−H), 6.99−6.83 (m, 2H, Ar−H and Ph-H), 4.51 and 3.61 (dd, 3JHH
= 10.8 Hz, 3.6 Hz, 1H, CH), 4.35−4.09 (m, 5H, CH and O−CH2-
CH2-O), 3.03−2.89 (m, 1H, CH2), 2.76−2.60 (m, 1H, CH2), 1.53−
1.43 (m, 3H, CH3); 13C NMR (100 MHz, DMSO-d6) δ 173.8 and
168.8, 146.8, 145.2 and 145.1, 143.6 and 143.6, 143.2, 137.7 and 137.1,
135.9 and 135.8, 127.7 and 127.5, 126.9 and 126.8, 120.7 and 120.7,
120.6 and 120.3, 118.5 and 118.4, 117.6 and 117.5, 115.3 and 114.9,
111.1 and 111.0, 106.2 and 105.9, 64.3 and 64.3, 64.1 and 64.0, 56.6
and 53.6, 48.5 and 47.9, 25.5 and 25.0, 19.9 and 19.6; HRMS (ESI)
calcd for C22H23N4O3 [M + H]+ 391.1765, found 391.1763.
1H, Py-H), 8.36 and 8.10 (s, 1H, NCH), 7.95 and 7.80 (d, JHH
=
3
4
8.0 Hz, 1H, Py-H), 7.88 and 7.75 (td, JHH = 7.6 Hz, JHH = 1.2 Hz,
1H, Py-H), 7.44−7.33 (m, 2H, Py-H and Ar−H), 7.31 and 7.30 (d,
3
3JHH = 7.6 Hz, 1H, Ar−H), 7.03 (t, JHH = 7.2 Hz, 1H, Ar−H), 6.95
and 6.92 (t, 3JHH = 7.2 Hz, 1H, Ar−H), 4.50 and 3.63 (dd, 3JHH = 10.8
3
Hz, 4.0 Hz, 1H, CH), 4.16 (q, JHH = 8.0 Hz, 1H, CH), 2.94 (ddd,
2JHH = 14.8 Hz, 3JHH = 4.4 Hz, 4JHH = 1.6 Hz, 1H, CH2), 2.72 and 2.63
2
3
4
(ddd, JHH = 14.8 Hz, JHH = 10.8 Hz, JHH = 2.0 Hz, 1H, CH2), 1.46
and 1.45 (d, 3JHH = 6.8 Hz, 3H, CH3); 13C NMR (100 MHz, DMSO-
d6) δ 174.8 and 169.5, 153.4 and 153.0, 149.5, 147.2 and 143.9, 137.8
and 137.8, 136.9 and 136.9, 135.8 and 135.8, 126.9 and 126.9, 124.4
and 124.2, 120.6, 119.9 and 119.6, 118.4, 117.5 and 117.5, 111.0, 106.1
and 106.0, 56.8 and 53.2, 48.5 and 47.7, 25.5, 19.9; HRMS (ESI) calcd
for C19H20N5O [M + H]+ 334.1662, found 334.1666.
Data for N′-((6-Hydroxynaphthalen-2-yl)methylene)-(3S)-1-
methyl-2,3,4,9-tetrahydro-1H-pyrido[3,4-b]indole-3-carbohydra-
zide (18). This compound needed recrystallization from methanol to
removed the remaining aldehyde. This compound was obtained as
1
yellow solid in 70% yield: mp = 275−278 °C; H NMR (400 MHz,
Data for N′-(Furan-2-ylmethylene)-(3S)-1-methyl-2,3,4,9-tetrahy-
dro-1H-pyrido[3,4-b]indole-3-carbohydrazide (22). The crude prod-
uct was chromatographed on silica gel (DCM/MeOH = 40:1 → 30:1
→ 20:1) to give two compounds. Compound 22a was obtained as
DMSO-d6) δ 11.46 and 11.35 (s, 1H, NH), 10.85 and 10.81 (s, 1H,
OC-NH), 9.98 (s, 1H, OH), 8.42 and 8.16 (s, 1H, NCH), 7.99
3
and 7.91 (s, 1H, Naphthalene-H), 7.85 (d, JHH = 9.2 Hz, 1H,
1
3
yellow solid in 55% yield: mp = 144−148 °C; H NMR (400 MHz,
Naphthalene-H), 7.80 and 7.61 (d, JHH = 8.8 Hz, 1H, Naphthalene-
3
CDCl3) δ 10.33 and 10.20 (s, 1H, NH), 8.26 and 8.11 (s, 1H, OC-
H), 7.74 and 7.70 (d, JHH = 8.8 Hz, 1H, Naphthalene-H), 7.42 and
3
NH), 8.04 and 7.72 (s, 1H, NCH), 7.51−7.38 (m, 2H, Ar−H and
7.37 (d, JHH = 7.6 Hz, 1H, Ar−H), 7.16 and 7.08 (s, 1H,
furan-H), 7.34 and 7.31 (d, 3JHH = 7.6 Hz, 1H, Ar−H), 7.14 (t, 3JHH
=
3
Naphthalene-H), 7.15−7.09 (m, 1H, Naphthalene-H), 7.04 (t, JHH
6.8 Hz, 1H, Ar−H), 7.08 (t, 3JHH = 7.2 Hz, 1H, Ar−H), 6.77 and 6.62
(s, 1H, furan-H), 6.45 and 6.42 (s, 1H, furan-H), 4.65 and 3.69 (d,
3JHH = 7.6 Hz, 1H, CH), 4.32−4.11 (m, 1H, CH), 3.30 and 3.19 (d,
2JHH = 13.6 Hz, 1H, CH2), 2.79 (t, 2JHH = 13.2 Hz, 1H, CH2), 1.97 (br,
1H, NH), 1.52 and 1.46 (d, 3JHH = 6.4 Hz, 3H, CH3); 13C NMR (100
MHz, DMSO-d6) δ 174.4 and 169.1, 149.5 and 149.1, 145.1 and 144.9,
137.8 and 137.7, 136.9, 135.8, 126.9 and 126.9, 120.6, 118.4, 117.5,
113.6 and 113.3, 112.2 and 112.1, 111.0, 106.2 and 106.1, 56.7 and
53.2, 48.5 and 47.7, 25.5 and 25.4, 19.9; HRMS (ESI) calcd for
C18H19N4O2 [M + H]+ 323.1503, found 323.1505.
= 7.6 Hz, 1H, Ar−H), 7.00−6.90 (m, 1H, Ar−H), 4.52 and 3.63 (d,
3JHH = 8.4 Hz, 1H, CH), 4.25−4.10 (m, 1H, CH), 3.02−2.90 (m, 1H,
CH2), 2.73 and 2.65 (t, 2JHH = 12.8 Hz, 1H, CH2), 1.47 (d, 3JHH = 6.0
Hz, 3H, CH3); 13C NMR (100 MHz, DMSO-d6) δ 174.5 and 169.0,
156.5 and 156.5, 147.4 and 144.0, 137.9, 135.8 and 135.8, 135.6 and
135.4, 130.1 and 130.0, 129.0, 128.7 and 128.7, 127.3 and 127.3, 127.0
and 126.9, 126.9 and 126.8, 123.0 and 122.3, 120.6, 119.2 and 119.2,
118.4, 117.5, 111.0, 109.1, 106.1 and 106.0, 56.7 and 53.3, 48.4 and
47.7, 25.6 and 25.4, 20.0 and 20.0; HRMS (ESI) calcd for C24H23N4O2
[M + H]+ 399.1816, found 399.1822.
Compound 22b was obtained as yellow solid in 3% yield: mp =
154−157 °C; 1H NMR (400 MHz, CDCl3) δ 12.00 and 10.44 (s, 1H,
NH), 8.00 and 7.94 (s, 1H, OC-NH), 7.64 and 7.60 (s, 1H, N
CH), 7.52 and 7.46 (d, 3JHH = 7.6 Hz, 1H, Ar−H), 7.40−7.30 (m, 2H,
Data for N′-(Pyridin-4-ylmethylene)-(3S)-1-methyl-2,3,4,9-tetra-
hydro-1H-pyrido[3,4-b]indole-3-carbohydrazide (19). This com-
pound was obtained as yellow solid in 79% yield: mp = 235−239
°C; 1H NMR (400 MHz, DMSO-d6) δ 11.84 and 11.69 (s, 1H, NH),
10.90 and 10.84 (s, 1H, OC-NH), 8.65 and 8.57 (d, 3JHH = 5.6 Hz,
3
Ar−H and furan-H), 7.16 (t, 3JHH = 7.2 Hz, 1H, Ar−H), 7.11 (t, JHH
D
dx.doi.org/10.1021/jf503794g | J. Agric. Food Chem. XXXX, XXX, XXX−XXX

Journal of Agricultural and Food Chemistry
Article
= 7.2 Hz, 1H, Ar−H), 6.78 and 6.70 (s, 1H, furan-H), 6.60 and 6.55 (s,
135.9, 128.5 and 128.1, 127.3 and 127.2, 121.8 and 121.7, 119.6 and
119.5, 118.3 and 118.1, 111.1, 107.9 and 107.7, 57.2 and 54.1, 49.4 and
48.0, 25.7 and 25.2, 20.2 and 20.2, 18.7 and 18.6; HRMS (ESI) calcd
for C18H20N5O [M + H]+ 297.1710, found 297.1714.
3
1H, furan-H), 4.67 and 3.82 (d, JHH = 8.8 Hz, 1H, CH), 4.36−4.25
(m, 1H, CH), 3.30 and 3.20 (d, 2JHH = 15.2 Hz, 1H, CH2), 2.92−2.75
(m, 1H, CH2), 1.72 (br, 1H, NH), 1.61−1.49 (m, 3H, CH3); 13C
NMR (100 MHz, DMSO-d6) δ 169.4, 147.8, 145.6, 138.2, 135.8,
130.8, 127.0, 120.6, 118.4, 117.6, 116.2, 112.5, 111.0, 106.8, 56.9, 48.8,
24.4, 19.9; HRMS (ESI) calcd for C18H19N4O2 [M + H]+ 323.1503,
found 323.1509.
Synthesis of N′-(Butylidene)-(3S)-1-methyl-2,3,4,9-tetrahy-
dro-1H-pyrido[3,4-b]indole-3-carbohydrazide (27).19 To a sol-
ution of compound 1 (0.50 g, 2.05 mmol) in THF (35 mL) was added
dropwise n-butanal (0.30 g, 4.10 mmol) at room temperature, After 3
h, TLC analysis showed the complete consumption of compound 1.
Then the mixture was concentrated in vacuo, and the crude product
was purified by column chromatography on silica gel eluted with
DCM/MeOH (25:1 → 10:1) to give 27 as a yellow solid in 71% yield:
Data for N′-((1H-Pyrrol-2-yl)methylene)-(3S)-1-methyl-2,3,4,9-
tetrahydro-1H-pyrido[3,4-b]indole-3-carbohydrazide (23). This
compound was obtained as red solid in 75% yield: mp = 207−209
°C; 1H NMR (400 MHz, DMSO-d6) δ 11.52 and 11.31 (s, 1H, NH),
11.16 and 11.05 (s, 1H, Pyrrole-NH), 10.83 and 10.80 (s, 1H, OC-
NH), 8.15 and 7.88 (s, 1H, NCH), 7.45−7.35 (m, 1H, Ar−H), 7.30
(d, 3JHH = 7.2 Hz, 1H, Ar−H), 7.03 (t, 3JHH = 7.2 Hz, 1H, Ar−H), 6.95
1
mp = 113−117 °C; H NMR (400 MHz, DMSO-d6) δ 11.04 and
10.97 (s, 1H, NH), 10.82 and 10.80 (s, 1H, OC-NH), 7.59 and
3
7.42−7.33 (m, 2H, NCH and Ar−H), 7.30 (d, JHH = 7.6 Hz, 1H,
3
Ph-H), 7.03 (t, 3JHH = 6.8 Hz, 1H, Ar−H), 6.95 (t, 3JHH = 6.8 Hz, 1H,
Ar−H), 4.28 and 3.51 (d, 3JHH = 9.6 Hz, 1H, CH), 4.19−4.04 (m, 1H,
CH), 2.89 (d, 2JHH = 14.0 Hz, 1H, CH2), 2.67 and 2.57 (t, 2JHH = 12.8
Hz, 1H, CH2), 2.25−2.10 (m, 2H, CH2), 1.55−1.40 (m, 5H, CH3 and
(t, JHH = 7.2 Hz, 1H, Ar−H), 6.90 and 6.83 (s, 1H, Pyrrole-H), 6.45
and 6.39 (s, 1H, Pyrrole-H), 6.13 and 6.08 (s, 1H, Pyrrole-H), 4.60
and 3.58 (d, 3JHH = 8.8 Hz, 1H, CH), 4.24−4.08 (m, 1H, CH), 2.98−
2.88 (m, 1H, CH2), 2.70−2.59 (m, 1H, CH2), 1.50−1.45 (m, 3H,
CH3); 13C NMR (100 MHz, DMSO-d6) δ 173.9 and 168.4, 140.2,
137.8, 136.4, 135.8, 127.0 and 126.9, 121.9, 120.6 and 120.6, 118.4,
117.7 and 117.5, 113.3 and 112.8, 111.0, 109.2 and 109.1, 106.4 and
106.2, 56.6 and 52.6, 48.4 and 47.7, 25.6 and 25.1, 19.9 and 19.7;
HRMS (ESI) calcd for C18H20N5O [M + H]+ 322.1662, found
322.1668.
3
CH2), 0.92 and 0.86 (t, JHH = 6.8 Hz, 3H, CH3); 13C NMR (100
MHz, DMSO-d6) δ 173.9 and 168.7, 151.3 and 147.8, 137.8 and 137.7,
135.9, 126.9, 120.6, 118.4, 117.5 and 117.5, 111.0, 106.2 and 106.1,
56.5 and 53.3, 48.4 and 47.7, 33.9 and 33.7, 25.5 and 25.2, 20.0, 19.5
and 19.2, 13.7 and 13.5; HRMS (ESI) calcd for C17H23N4O [M + H]+
299.1866, found 299.1870.
Data for N′-(Thiophen-2-ylmethylene)-(3S)-1-methyl-2,3,4,9-tet-
rahydro-1H-pyrido[3,4-b]indole-3-carbohydrazide (24). This com-
pound was obtained as yellow solid in 76% yield: mp = 139−141 °C;
1H NMR (400 MHz, DMSO-d6) δ 11.55 and 11.39 (s, 1H, NH),
Compounds 28 and 29 were synthesized using a similar procedure
as compound 27.
Data for N′-(Octylidene)-(3S)-1-methyl-2,3,4,9-tetrahydro-1H-
pyrido[3,4-b]indole-3-carbohydrazide (28). The crude product was
purified by column chromatography on silica gel eluted with DCM/
MeOH (40:1 → 30:1 → 10:1) to give 28 as a yellow solid in 78%
yield: mp = 68−71 °C; 1H NMR (400 MHz, CDCl3) δ 9.94 and 9.02
10.90 and 10.83 (s, 1H, OC-NH), 8.56 and 8.24 (s, 1H, NCH),
3
7.67 and 7.54 (d, JHH = 4.8 Hz, 1H, Thiophene-H), 7.47−7.35 (m,
2H, Thiophene-H and Ar−H), 7.35−7.28 (m, 1H, Ar−H), 7.14 and
7.09 (t, 3JHH = 4.4 Hz, 1H, Thiophene-H), 7.04 (t, 3JHH = 7.2 Hz, 1H,
Ar−H), 7.00−6.90 (m, 1H, Ar−H), 4.38 and 3.62 (dd, 3JHH = 8.8 Hz,
4.0 Hz, 1H, CH), 4.29−4.10 (m, 1H, CH), 3.04−2.89 (m, 1H, CH2),
2.77−2.61 (m, 1H, CH2), 1.54−1.38 (m, 3H, CH3); 13C NMR (100
MHz, DMSO-d6) δ 173.6 and 168.9, 142.3, 139.1 and 138.9, 138.6 and
137.6, 135.9 and 135.8, 130.8 and 130.4, 128.8 and 128.4, 128.0 and
127.9, 126.9 and 126.8, 120.7 and 120.6, 118.5 and 118.4, 117.5, 111.1
and 111.0, 106.1 and 105.9, 56.7 and 53.7, 48.5 and 47.9, 25.4 and
25.0, 19.8 and 19.6; HRMS (ESI) calcd for C18H19N4OS [M + H]+
339.1274, found 339.1278.
(s, 1H, NH), 7.90 and 7.50 (s, 1H, NCH), 7.50 and 7.46 (d, 3JHH
=
8.0 Hz, 1H, Ar−H), 7.34 and 7.32 (d, 3JHH = 8.4 Hz, 1H, Ar−H), 7.17
(t, 3JHH = 7.2 Hz, 1H, Ar−H), 7.11 (t, 3JHH = 7.2 Hz, 1H, Ar−H), 4.53
3
and 3.70 (dd, JHH = 11.2 Hz, 4.4 Hz, 1H, CH), 4.33−4.17 (m, 1H,
2
3
CH), 3.37 and 3.13 (dd, JHH = 14.4 Hz, JHH = 2.8 Hz, 1H, CH2),
2.83−2.72 (m, 1H, CH2), 2.38 and 2.22 (q, 3JHH = 7.2 Hz, 2H, CH2),
1.96 (br, 1H, NH), 1.58−1.43 (m, 4H, CH2 and CH3), 1.42−1.18 (m,
9H, CH2), 0.95−0.81 (m, 3H, CH3); 13C NMR (100 MHz, CDCl3) δ
175.2 and 168.6, 152.8 and 148.8, 136.7 and 136.6, 135.9 and 135.9,
127.3 and 127.2, 122.0 and 121.8, 119.7 and 119.5, 118.4 and 118.1,
111.0, 108.4 and 107.9, 57.3 and 54.1, 49.5 and 47.9, 32.4 and 32.1,
31.7 and 31.7, 29.3, 29.1 and 29.0, 26.7 and 26.3, 25.6 and 25.1, 22.6
and 22.6, 20.2, 14.1 and 14.1; HRMS (ESI) calcd for C21H31N4O [M +
H]+ 355.2493, found 355.2492.
Data for N′-((1H-imidazol-2-yl)methylene)-(3S)-1-methyl-2,3,4,9-
tetrahydro-1H-pyrido[3,4-b]indole-3-carbohydrazide (25). This
compound was obtained as green solid in 81% yield: mp = 188−
1
190 °C; H NMR (400 MHz, DMSO-d6) δ 14.10, 13.33, 12.81, and
12.57 (s, 1H, Imidzole-NH), 12.93 and 11.47 (s, 1H, NH), 10.84 and
Data for N′-(Cyclohexylmethylene)-(3S)-1-methyl-2,3,4,9-tetra-
hydro-1H-pyrido[3,4-b]indole-3-carbohydrazide (29). The crude
product was purified by column chromatography on silica gel eluted
with DCM/MeOH (100:1 → 40:1 → 20:1) to give 29 as a yellow
solid in 92% yield: mp = 123−126 °C; 1H NMR (400 MHz, CDCl3) δ
9.88 and 9.21 (s, 1H, NH), 7.98 and 7.96 (s, 1H, NCH), 7.49 and
7.47 (d, 3JHH = 8.0 Hz, 1H, Ar−H), 7.38−7.29 and 7.05 (m, 2H, Ar−
H and OC-NH), 7.16 and 7.16 (t, 3JHH = 7.2 Hz, 1H, Ar−H), 7.10
10.81 (s, 1H, OC-NH), 8.45, 8.26, 7.95, and 7.44 (s, 1H, NCH),
7.42−6.88 (m, 6H, Ar−H and Imidzole-H), 4.65, 4.50, 3.69, and 3.63
2
(m, 1H, CH), 4.37 and 4.17 (m, 1H, CH), 3.03 and 2.95 (d, JHH
=
14.0 Hz, 1H, CH2), 2.78−2.56 (m, 1H, CH2), 1.52−1.42 (m, 3H,
CH3); 13C NMR (100 MHz, DMSO-d6) δ 174.9, 174.7, 170.4 and
169.2, 142.6, 142.5, 141.1 and 141.0, 139.0, 138.2, 137.8 and 137.7,
135.8 and 135.2, 129.6 and 129.3, 127.0, 126.9, 126.9 and 126.8, 120.6
and 120.6, 118.4, 117.7 and 117.5, 111.0 and 111.0, 106.5, 106.4, 106.2
and 106.0, 57.2, 56.8, 53.1 and 52.5, 48.7, 48.5, 47.9 and 47.7, 25.6,
25.3, 25.3 and 25.1, 20.0, 20.0, 19.8 and 19.8; HRMS (ESI) calcd for
C17H19N6O [M + H]+ 323.1615, found 323.1620.
3
3
(t, JHH = 7.2 Hz, 1H, Ar−H), 4.52 and 3.66 (dd, JHH = 10.8 Hz, 4.4
Hz, 1H, CH), 4.28 and 4.18 (q, 3JHH = 6.8 Hz, 1H, CH), 3.35 and 3.12
2
3
(dd, JHH = 15.6 Hz, JHH = 2.8 Hz, 1H, CH2), 2.85−2.72 (m, 1H,
CH2), 2.47−2.35 and 2.23−2.13 (m, 1H, CH), 1.94 (br, 1H, NH),
1.87−1.58 (m, 4H, CH2), 1.53 and 1.48 (d, 3JHH = 10.8 Hz, 3H, CH3),
1.38−1.11 (m, 6H, CH2), 0.95−0.81 (m, 3H, CH3); 13C NMR (100
MHz, CDCl3) δ 175.3 and 168.6, 156.3 and 152.2, 136.9 and 136.6,
135.9 and 135.8, 127.4 and 127.3, 122.0 and 121.7, 119.8 and 119.5,
118.4 and 118.1, 110.9, 108.5 and 108.0, 57.4 and 54.2, 49.5 and 47.9,
40.8 and 40.4, 30.2, 30.0 and 30.0, 25.9 and 25.8, 25.6 and 25.4, 25.3
and 25.1, 20.3 and 20.3; HRMS (ESI) calcd for C20H27N4O [M + H]+
339.2180, found 339.2179.
Data for (N′-((E)-But-2-enylidene)-(3S)-1-methyl-2,3,4,9-tetrahy-
dro-1H-pyrido[3,4-b]indole-3-carbohydrazide (26). The crude pro-
dut was chromatographed on silica gel to give 26 as a yellow solid in
1
79% yield: mp = 145−148 °C; H NMR (400 MHz, CDCl3) δ 10.10
and 9.63 (s, 1H, NH), 8.23 and 8.15 (s, 1H, OC-NH), 7.72 and
7.37 (d, 3JHH = 8.4 Hz, 1H, NCH), 7.43 (d, 3JHH = 6.8 Hz, 1H, Ar−
H), 7.35−7.27 (m, 1H, Ar−H), 7.20−7.00 (m, 2H, Ar−H), 6.42−5.94
(m, 2H, CHCH), 4.57−4.48 and 3.70−3.58 (m, 1H, CH), 4.31−4.06
(m, 1H, CH), 3.27 and 3.12 (d, 2JHH = 14.4 Hz, 1H, CH2), 2.85−2.66
(m, 1H, CH2), 2.09 (br, 1H, NH), 1.92−1.75 (m, 3H, CH3), 1.50 and
Synthesis of N′-(2,2-Dimethylpropylidene)-(3S)-1-methyl-
2,3,4,9-tetrahydro-1H-pyrido[3,4-b]indole-3-carbohydrazide
(30).19 To a solution of compound 1 (0.38 g, 1.56 mmol) in THF (35
mL) was added dropwise trimethylacetaldehyde (0.27 g, 3.11 mmol)
3
1.43 (d, JHH = 5.6 Hz, 3H, CH3); 13C NMR (100 MHz, CDCl3) δ
175.5 and 169.2, 150.3 and 147.2, 139.7 and 138.9, 136.8, 136.0 and
E
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Scheme 1. Synthesis of Compound 1
Scheme 2. Synthesis of Compounds 3 and 5−33
at room temperature, After 6 h, TLC analysis showed the complete
consumption of compound 1. Then the mixture was concentrated in
vacuo and dried without further operation to give 30 as a yellow solid
in 95% yield: mp = 140−141 °C; 1H NMR (100 MHz, CDCl3) δ 9.93
and 9.39 (s, 1H, NH), 8.13 and 8.09 (s, 1H, NCH), 7.53−7.02 (m,
4H, Ar−H), 4.51 and 3.67 (d, 3JHH = 8.4 Hz, 1H, CH), 4.37−4.16 (m,
(m, 1H, CH2), 2.76−2.59 (m, 1H, CH2), 3.31 and 3.30 (s, 3H, CH3),
1.46 (d, 3JHH = 6.4 Hz, 3H, CH3); 13C NMR (100 MHz, DMSO-d6) δ
175.3 and 169.2, 151.9 and 148.0, 138.1, 137.9 and 137.6, 135.8 and
135.8, 129.3 and 129.1, 128.4 and 128.4, 126.9, 126.4 and 125.9, 120.6,
118.4, 117.5 and 117.5, 111.0, 106.4 and 106.1, 56.5 and 53.7, 48.4 and
47.7, 25.5 and 25.3, 19.9, 13.7 and 13.4; HRMS (ESI) calcd for
C21H23N4O [M + H]+ 347.1867, found 347.1872.
1H, CH), 3.33 and 3.13 (d, 2JHH = 14.4 Hz, 1H, CH2), 2.79 (t, 2JHH
=
13.2 Hz, 1H, CH2), 2.09 (br, 1H, NH), 1.58−1.41 (m, 3H, CH3), 1.15
and 1.06 (s, 9H, CH3); 13C NMR (100 MHz, CDCl3) δ 176.0 and
169.3, 160.9 and 156.5, 137.3, 136.7 and 136.6, 128.1 and 127.9, 122.6
and 122.4, 120.4 and 120.2, 119.1 and 118.8, 111.7, 109.0 and 108.5,
58.1 and 55.1, 50.2 and 48.7, 35.8 and 35.6, 31.0 and 27.9, 28.1 and
28.0, 26.1 and 25.8, 20.9; HRMS (ESI) calcd for C18H25N4O [M +
H]+ 313.2023, found 313.2028.
Compounds 32 and 33 were synthesized using a similar procedure
as compound 31.
Data for N′-(3,3-Dimethylbutan-2-ylidene)-(3S)-1-methyl-2,3,4,9-
tetrahydro-1H-pyrido[3,4-b]indole-3-carbohydrazide (32). The
crude product was purified by column chromatography on silica gel
eluted with DCM/MeOH (30:1 → 10:1) to give 32 as a yellow solid
1
in 63% yield: mp = 103−107 °C; H NMR (400 MHz, DMSO-d6) δ
Synthesis of N′-(1-Phenylethylidene)-(3S)-1-methyl-2,3,4,9-
tetrahydro-1H-pyrido[3,4-b]indole-3-carbohydrazide (31).20
To a solution of compound 1 (0.50 g, 2.05 mmol) in EtOH (35
mL) was added acetophenone (0.27 g, 2.25 mmol), and then the
reaction mixture was heated to reflux. After 6 h, the reaction was
complete detected by TLC. The mixture was concentrated in vacuo,
and the crude product was purified by column chromatography on
silica gel eluted with DCM/MeOH (30:1 → 10:1) to give 31 as a light
10.80 and 10.79 (s, 1H, N−H), 10.18 and 10.02 (s, 1H, OC-
NH),7.40 and 7.35 (d, 3JHH = 7.6 Hz, 1H, Ar−H), 7.29 (d, 3JHH = 8.0
Hz, 1H, Ar−H), 7.03 (t, 3JHH = 7.2 Hz, 1H, Ar−H), 6.95 and 6.93 (t,
3JHH = 6.8 Hz, 1H, Ar−H), 4.26 and 3.65 (dd, 3JHH = 10.8 Hz, 4.0 Hz,
2
3
1H, CH), 4.16−4.07 (m, 1H, CH), 2.94 (dd, JHH = 14.8 Hz, JHH
=
2.8 Hz, 1H, CH2), 2.70−2.53 (m, 1H, CH2), 1.87 and 1.85 (s, 3H,
CH3), 1.43 (d, 3JHH = 6.4 Hz, 3H, CH3), 1.12 and 1.04 (s, 9H, CH3);
13C NMR (100 MHz, DMSO-d6) δ 174.7 and 168.6, 163.2 and 158.5,
137.9 and 137.5, 135.8, 126.9, 120.6 and 120.5, 118.4, 117.5 and 117.4,
111.0, 106.5 and 106.2, 56.6 and 54.4, 48.4 and 47.8, 38.5 and 38.3,
27.6 and 27.5, 26.0, 25.3 and 24.8, 20.0 and 19.9, 12.0 and 11.8;
1
yellow solid in 56% yield: mp = 221−224 °C; H NMR (400 MHz,
DMSO-d6) δ 10.84 and 10.81 (s, 1H, N−H), 10.69 and 10.48 (s, 1H,
OC-NH),7.89−7.77 and 7.76−7.67 (m, 2H, Ph-H), 7.48−7.27 (m,
5H, Ph-H and Ar−H), 7.08−6.88 (m, 2H, Ar−H), 4.53 and 3.78 (dd,
3JHH = 10.8 Hz, 3.6 Hz, 1H, CH), 4.23−4.11 (m, 1H, CH), 3.03−2.93
F
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HRMS (ESI) calcd for C19H27N4O [M + H]+ 327.2180, found
327.2186.
Scheme 4. Prediction of the Configuration
Data for N′-Cyclohexylidene-(3S)-1-methyl-2,3,4,9-tetrahydro-
1H-pyrido[3,4-b]indole-3-carbohydrazide (33). The crude product
was purified by column chromatography on silica gel eluted with
DCM/MeOH (30:1 → 10:1) to give 33 as a yellow solid in 60% yield:
1
mp = 131−135 °C; H NMR (400 MHz, DMSO-d6) δ 10.81 and
10.78 (s, 1H, NH), 10.39 and 10.24 (s, 1H, OC-NH),7.39 and 7.36
3
3
(d, JHH = 8.0 Hz, 1H, Ar−H), 7.29 (d, JHH = 7.6 Hz, 1H, Ar−H),
7.02 (t, 3JHH = 7.2 Hz, 1H, Ar−H), 6.94 (t, 3JHH = 7.2 Hz, 1H, Ar−H),
4.36 and 3.62 (dd, JHH = 10.0 Hz, 3.6 Hz, 1H, CH), 4.18−4.04 (m,
3
1H, CH), 2.94−2.84 (m, 1H, CH2), 2.69−2.58 (m, 1H, CH2), 2.40−
2.12 (m, 4H, CH2), 1.71−1.51 (m, 6H, CH2), 1.42 (d, 3JHH = 6.0 Hz,
3H, CH3); 13C NMR (100 MHz, DMSO-d6) δ 168.7, 161.5, 137.8,
135.8, 126.9 and 126.8, 120.6 and 120.5, 118.4, 117.5, 111.0, 106.3,
56.3 and 53.3, 48.3 and 47.7, 35.2 and 35.0, 27.0 and 26.8, 25.7 and
25.5, 25.1, 19.9 and 19.8; HRMS (ESI) calcd for C19H25N4O [M +
H]+ 325.2023, found 325.2023.
Biological Assay. The anti-TMV and fungicidal and insecticidal
activities of the synthesized compounds were tested using our
previously reported methods.21,22
Field Trials. The experiments were carried out between June and
July in 2014. The trial against TMV was performed in the trial area of
Institute of Environment and Resources, Yunnan Provincial Academy
of Agricultural Sciences, Kunming, Yunnan Province, China. The
experimental design was a randomized complete block with three
treatments. The untreated plots served as a blank control. The
formulation of compounds 9 (1% EC) and 29 (1% ME) was prepared
in our lab. They were diluted to 10, 50, or 100 g(ai)/ha (weight
(gram) of active ingredient/hectare) before use. The formulation of
moroxydine hydrochloride−cupric acetate (20% WP) and amino-
oligosaccharins (5% aqueous solution) were obtained from Wansheng
Biological Pesticide Co., Ltd. (Shandong, China) and Hainan zhengye
zhong nong Co., Ltd. (Hainan, China) respectively. The incidence was
surveyed 10 days after the third time spraying. The severity of the virus
disease was divided into 0−4 grade. The disease index and control
effect can be calculated by using the following formulas:
Figure 3. Compound 21.
reacted with thionyl chloride and methanol to form methyl
ester, which subsequently reacted with hydrazine hydrate to
give compound 1.
The compounds 3 and 6−33 containing the acylhydrazone
fragment all can be synthesized from lead compound 1 by
reaction with the corresponding aldehydes or ketones under
different reaction conditions (Scheme 2). To be specific, when
aromatic aldehydes were used as starting materials, the
corresponding reactions were carried out in toluene under
reflux conditions. When the reaction was complete, the reaction
system was cooled to room temperature, and the solid products
precipitated because of their poor solubility in toluene. By this
approach, compounds 3 and 6−26 could be easily obtained in
60%−91% yields. When aliphatic aldehydes were used as
starting materials, the corresponding reactions were carried out
in tetrahydrofuran at room temperature. When the reaction was
complete, compounds 27−30 could be obtained by column
chromatography in 71%−95% yields. When ketones were used,
these reactions were carried out in ethanol under reflux
conditions. When the reaction was complete, compounds 31−
33 could be obtained by column chromatography in 56%−63%
yields.
disease index
I% = [
(N n)/(N4)] × 100
∑
n
′
control effect (%) = 1 − (IbcI)/(IbcI′) × 100
where Nn is the number of diseased plants of each relative level values,
n is the relative level values (n = 0−4), N is the total number of
diseased plants, I′bc is the disease index of the blank control area before
spraying, Ibc is the disease index of the blank control area 10 days after
the third time spraying, I′ is the disease index of the spraying area
before spraying; and I is the disease index of the spraying area 10 days
after the third time spraying.
RESULTS AND DISCUSSION
■
Synthesis. The synthesis of the lead compound 1 is shown
in Scheme 1. Using L-tryptophan as starting material, the
tetrahydrocarboline skeleton of 1 was constructed through
Pictet−Spengler cyclization, and then the given carboxylic acid
In addition, 1-methyl-9H-carbazole-3-carboxylic acid hydra-
zide 2 could be prepared from ethyl 1-methyl-9H-pyrido[3,4-
b]indole-3-carboxylate, which was synthesized from tryptophan
through a literature protocol.17 Compound 2 reacted with
benzaldehyde using a similar procedure as the preparation of
compound 3 to give 4 in 91% yield (Scheme 3).
Scheme 3. Synthesis of Compounds 2 and 4
Configuration. At room temperature, the 1H and 13C NMR
spectra of most of target compounds exhibited two sets of
signals, which aroused our attention. Initially, this phenomenon
was thought to be due to the presence of cis−trans isomers
1
(prediction A, Scheme 4); however, in the H NMR spectrum
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1
Figure 4. H NMR of Compound 21 at Different Temperatures.
of 4 only one set of signals was observed, which suggested such
phenomenon resulted from other reasons. Meanwhile, the
presence of diastereoisomers (prediction B, Scheme 4) as a
possible reason was excluded (the 1H NMR spectra of
precursor 1 as well as compound 21 (Figures 3, 4 and 5)
clearly showed the presence of diastereoisomeric mixtures, and
the ratio is 10:1). Eventually, hydrogen-bonding effects
(prediction C, Scheme 4) was put forward, and variable-
temperature (VT) H NMR studies (Figures 4 and 5) of
compound 21 were performed in order to confirm this
1
H
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1
Figure 5. H NMR (H2) of compounds 1 and 21.
Table 1. In Vitro Antiviral Activities of Compounds 1−33
against TMV
Figure 6. Hydrogen-bonding effects of compound 25.
hypothesis. As we can see, the coalescence of signals was
1
observed in the entire H NMR spectrum with the rise of the
temperature (20 → 40 → 60 → 80 °C). Therefore, the VT 1H
1
NMR studies indicated that the doubling of signals in the H
and 13C NMR spectra was very likely due to hydrogen-bonding
effects. In addition, it also could explain the appearance of four
sets of signals in compound 25 (Figure 6).
Antiviral Activities. In our previous work, high antiviral
lead compound 1 was found, on the basis of which, a series of
derivatives containing acylhydrazone structure were synthesized
to investigate the influence of the variation in the basic skeleton
of 1. The acylhydrazone compounds exhibited excellent
antiviral activity. The antiviral results in vitro and in vivo
(protection, inactivation, and curative) are listed in Tables 1
and 2. Overall, compared with ribavirin, most of the newly
designed compounds exhibited desirable anti-TMV activity in
vitro. Compounds 8, 9, 12, 16, 28, 29, and 30 showed much
higher inhibition than that of ribavirin and their precursor 1 at
500 and 100 μg/mL. Especially, the activity of compound 16
was 36.9% at 100 μg/mL, which was very close to that of
Ribavirin (40%) at 500 μg/mL.
When the C ring (Figure 3) was tetrahydropyridine, the in
vitro activity of compound 1 (40.4%, 500 μg/mL) was a little
lower than that of the β-carboline compound 2 (43.9%, 500
μg/mL); however, activity of compound 3 (55.8%, 500 μg/
mL) was obviously higher than that of compound 4 (41.8%,
500 μg/mL). On the other hand, the activity of compound 1
(40.4%, 500 μg/mL) showed obviously lower inhibition than
that of compound 3 (55.8%, 500 μg/mL), which proved that it
is advantageous to improve the activity if the hydrazide
structure at the 3-position of tetrahydrogen-β-carboline was
changed to acylhydrazone. Hence, a representative set of
compounds containing acylhydrazone structure were designed
to search for better-activity compounds. Additionally, the
structure−activity relationship (SAR) demonstrated that the
chirality at the 3-position of tetrahydrogen-β-carboline affected
slightly the antiviral activity; for instance, compound 3 (3S,
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Table 2. In Vivo Antiviral Activities of Compounds 1−33 against TMV
55.8%, 500 μg/mL) was slightly more active than compound 5
(3R, 49.6%, 500 μg/mL).
Furthermore, it was noteworthy that the substituents of the
acylhydrazone moiety had a significant impact on anti-TMV
activity.
mL), the order of their activities was para > meta > ortho.
However, the activity was greatly reduced when the benzene
ring was changed to a naphthalene ring, such as 18 (40.9%, 500
μg/mL).
Interestingly, compound 16 (72.6%, 500 μg/mL) exhibited
excellent inhibitory effect; in contrast, compounds 15 (37.2%,
500 μg/mL) and 17 (46.5%, 500 μg/mL) were far less effective.
We speculated it could be ascribed to the tension on the five-
membered ring of 16, which inhibited the conjugation of the
lone-pair electrons of oxygen atoms with the benzene ring.
When R2 was hydrogen and R1 was a heterocyclic ring, the
anti-TMV activities of these compounds (19−25) reduced
obviously compared with those of compounds with substituted
benzene (3, 5−18).
When R2 was hydrogen and R1 was a benzene ring, it was
universally advantageous to anti-TMV activity if substituents
were introduced to the 4-position of the benzene ring such as 7
(56.8%, 500 μg/mL), 8 (62.8%, 500 μg/mL), 9 (67.3%, 500
μg/mL), and 12 (62.5%, 500 μg/mL). Monosubstitution or
multisubstitution on the benzene ring affected anti-TMV
activity to a certain extent, for instance, compared with
compounds 10 (56.8%, 500 μg/mL) and 11 (50.7%, 500 μg/
mL), the monosubstituted compound 9 exhibited significantly
higher activity. In addition, the position of the substituent
group also was favored to increase anti-TMV activity; the
activity of compound 12 (62.5%, 500 μg/mL) was much better
than that of 13 (53.2%, 500 μg/mL) and 14 (38.5%, 500 μg/
When R2 was hydrogen and R1 was an aliphatic substituent,
both the length of the aliphatic chain and the number of
branched chains affected anti-TMV activity; the more branched
and the longer was the aliphatic chain, the better was the
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Table 3. Fungicidal Activity of Compounds 1−4 and
Compounds 6−33 against Fourteen Kinds of
Phytopathogens
Table 4. Insecticidal Activity of Compounds 1−4 and
Compound 6−33 against Four Kinds of Insects
a
F.C., Fusarium oxysporum sp. cucumeris; C.H., Cercospora arachidicola
Hori; P.P., Physalospora piricola; A.S., Alternaria solani; F.G., Fusarium
graminearum; F.M., Fusarium moniliforme; S.S., Sclerotinia sclerotiorum;
P.C., Phytophthora capsici; R.C., Rhizoctonia cerealis; B.M., Bipolaris
maydis; W.A., watermelon anthracnose; P.I., Phytophthora infestans;
R.S., Rhizoctonia solani; B.C., Botrytis cinerea.
activity. For example, compound 26 containing an unsaturated
aliphatic chain (35.2%, 500 μg/mL) and 27 (32.6%, 500 μg/
mL) containing a saturated short-chain displayed lower anti-
TMV activities than 28 (61.3%, 500 μg/mL), 29 (59.6%, 500
μg/mL), and 30 (64.4%, 500 μg/mL). When R2 was methyl,
the compounds 31 (30.2%, 500 μg/mL), 32 (58.1%, 500 μg/
mL), and 33 (43.4%, 500 μg/mL) exhibited much lower
antiviral activities than 3 (55.8%, 500 μg/mL), 30 (64.4%, 500
μg/mL), and 29 (59.6%, 500 μg/mL), which meant that the
hydrogen-bond in NHCH played an important role in the
anti-TMV activity.
Further bioassay was conducted to investigate their
inactivation, curative, and protection effect in vivo (Table 2).
These results also showed that most of the compounds
exhibited excellent antiviral activity and similar structure−
activity relationship as the activities in vitro. Especially the
activities of compounds 9 (70.4%, 71.5%, and 64.2% at 500 μg/
mL), 12 (69.1%, 66.4%, and 70% at 500 μg/mL), 16 (74.5%,
69%, and 68.1% at 500 μg/mL), 28 (73.4%, 70.9%, and 59.6%
at 500 μg/mL), 29 (75.8%, 62.8%, and 69.2% at 500 μg/mL),
and 30 (63.7%, 68.2%, and 66.8% at 500 μg/mL) were much
superior to ribavirin (37.6%, 39.4%, and 37.9% at 500 μg/mL).
What is more, their anti-TMV activities (≥30%) at 100 μg/mL
were close to ribavirin at 500 μg/mL.
against 14 kinds of phytopathogens; especially compounds 6, 9,
and 11 exhibited desirable fungicidal activities against each of
the phytopathogens. Additionally, tetrahydrogen-β-carboline
derivatives exhibited higher activities than β-carboline deriva-
tives. When the skeleton was tetrahydrogen-β-carboline (C ring
was tetrahydropyridine), the fungicidal activities were greatly
affected by the substituents (R1 and R2) on the acylhydrozone.
The compounds with one hydrogen and one aryl group (3, 5−
25) exhibited high activities against fungi; whereas the
compounds with no hydrogen or with at least one alkyl
group (26−33) exhibited relatively poor activities. Note-
worthily, the compounds containing aryl groups exhibited
fungicidal activities to certain fungi selectively; they showed
apparently higher activities against Physalospora piricola,
Sclerotinia sclerotiorum, Phytophthora capsici, Rhizoctonia cerealis,
Bipolaris maydi, and Botrytis cinerea than the others.
Fungicidal Activity. The fungicidal results are listed in
Table 3. Most of β-carboline, tetrahydrogen-β-carboline
alkaloids, and their derivatives showed good fungicidal activities
K
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Journal of Agricultural and Food Chemistry
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compound 1 (40.4%, 40.0%, 42.3%, and 39.6% at 500 μg/
mL), and the relevant SAR was summarized. At the same time,
field trials of compounds 9 and 29 further demonstrated the
antiviral efficacy against TMV. In addition, we were pleased to
find that these compounds also showed broad-spectrum of
fungicidal activity and insecticidal activity. For instance,
compounds 3, 6, 9, and 11 exhibited good inhibition of the
14 kinds of phytopathogens; compounds 3 and 23 exhibited
moderate insecticidal activity against four kinds of investigated
insects. Further studies on structural optimization are in
progress in our laboratory.
Table 5. Results of Anti-TMV Activities of Field Trial in
2014 after 10 Days of Application of 9, 29, Amino-
oligosaccharins, and Moroxydine Hydrochloride−Cupric
Acetate
concn
(gai/ha)
a
b
c
d
compd
Ie′q
Ieq
E (%)
9 (1% EC)
10
50
1.14
1.86
0.00
1.10
0.79
0.00
2.56
3.33
1.52
4.92
0.00
1.52
0.00
0.00
5.23
3.33
65.28
80.82
100
10
100.00
70.91
29 (1% ME)
50
100.00
100.00
100
AUTHOR INFORMATION
■
blank control
Corresponding Author
*Qingmin Wang. Tel: +86-(0)22-23503952; Fax: +86-(0)22-
amino-oligosaccharins (5% aqueous
solution)
100
600
66.67
66.67
moroxydine hydrochloride−cupric
1.23
2.67
23503952; E-mail: wang98h@263.net; wangqm@nankai.edu.
cn.
acetate (20% WP)
a
EC, emulsifiable concentrate; ME, microemulsion; WP, wettable
b
Notes
powder. Ie′q, the average disease index of the spraying area before
spraying. Ieq, the average disease index of the spraying area 10 days
after the third time spraying. E, the average control effect 10 days
c
The authors declare no competing financial interest.
d
after the third time spraying.
ACKNOWLEDGMENTS
■
This work was supported by the National Natural Science
Foundation of China (Grants 21132003, 21121002, 21372131,
and 21102077), the Specialized Research Fund for the Doctoral
Program of Higher Education (Grant 20130031110017), and
the Science Technology Plan Project of China Tobacco
Yunnan Company (Grants 2012YN08, 2012YN11, and
2013YN37).
REFERENCES
■
(1) El-Wakeil, N. E. Botanical Pesticides and Their Mode of Action.
Gesunde Pflanz. 2013, 65, 125−149.
Figure 7. Effect of anti-TMV activities in field trials of compounds 9
and 29.
(2) Jacobson, M. Botanical pesticides, past, present, and future. ACS
Symp. Ser. 1989, 387, 1−10.
(3) Isman, M. B. Botanical insecticides: For richer, for poorer. Pest
Manage. Sci. 2008, 64 (1), 8−11.
Insecticidal Activity. These derivatives also showed broad-
spectrum insecticidal activities (Table 4). What is more,
compounds 3 and 23, which contain a phenyl group or a
pyrryl group, respectively, exhibited moderate insecticidal
activity against the four kinds of insects (Mythimna separata,
Helicoverpa armigera, Ostrinia nubilalis, and Culex pipiens
pallens).
Field Trials. Because compounds 9 and 29 exhibited
excellent anti-TMV activity in laboratory, 1% EC of compound
9 and 1% ME of compound 29 were evaluated for their anti-
TMV activities in Tengchong County, Yunnan Province, China.
Their efficacy was compared with the amino-oligosaccharins,
moroxydine hydrochloride−cupric acetate, and an untreated
control. Table 5 presents part of the results of field trials.
Compounds 9 and 29 (Figure 7) exhibited better activities than
control plant virus inhibitors. The results showed that the series
of compounds, especially compound 29, are promising for plant
virus control.
(4) Massaro, E. J. Handbook of Neurotoxicology; Humana Press:
Totowa, NJ, USA, 2002; p 237.
(5) Callaway, J. C.; McKenna, D. J.; Grob, C. S.; Brito, G. S.;
Raymon, L. P.; Poland, R. E.; Andrade, E. N.; Andrade, E. O.; Mash,
D. C. Pharmacokinetics of Hoasca alkaloids in healthy humans. J.
Ethnopharmacol. 1999, 65 (3), 243−256.
(6) Reniers, J.; Robert, S.; Frederick, R.; Masereel, B.; Vincent, S.;
Wouters, J. Synthesis and evaluation of β-carboline derivatives as
potential monoamine oxidase inhibitors. Bioorg. Med. Chem. 2011, 19
(1), 134−144.
(7) Srivastava, S. K.; Agarwal, A.; Chauhan, P. M. S.; Agarwal, S. K.;
Bhaduri, A. P.; Singh, S. N.; Fatima, N.; Chatterjee, R. K. Potent 1,3-
disubstituted-9H-pyrido[3,4-b]indoles as new lead compounds in
antifilarial chemotherapy. J. Med. Chem. 1999, 42 (9), 1667−1672.
(8) Tang, J. G.; Wang, Y. H.; Wang, R. R.; Dong, Z. J.; Yang, L. M.;
Zheng, Y. T.; Liu, J. K. Synthesis of analogues of flazin, in particular,
flazinamide, as promising anti-HIV agents. Chem. Biodiversity 2008, 5
(3), 447−460.
(9) Miller, J. F.; Turner, E. M.; Sherrill, R. G.; Gudmundsson, K.;
Sethna, A. S. P.; Brown, K. W.; Harvey, R.; Romines, K. R.; Golden, P.
Substituted tetrahydro-β-carbolines as potential agents for the
treatment of human papillomavirus infection. Bioorg. Med. Chem.
Lett. 2010, 20 (1), 256−259.
(10) Zhao, P. L.; Li, J.; Yang, G. F. Synthesis and insecticidal activity
of chromanone and chromone analogues of diacylhydrazines. Bioorg.
Med. Chem. 2007, 15, 1888−1895.
In summary, we designed and synthesized a series of novel
tetrahydrogen-β-carboline and β-carboline derivatives contain-
ing an acylhydrazide or acylhydrazone moiety at the 3-position.
The hydrogen bonding effect was verified via variable-
1
temperature (VT) H NMR studies. What is more, the target
compounds exhibited good anti-TMV activity in the laboratory;
especially, in vivo, inactivation, curative, and protection activities
of compounds 8, 9, 12, 16, 28, 29, and 30 were much higher
(57.1−75.8% at 500 μg/mL) than that of ribavirin (37.6%,
37.6%, 39.4%, and 37.9% at 500 μg/mL) and the lead
(11) Chen, Q.; Liu, Z. M.; Chen, C. N.; Jiang, L. L.; Yang, G. F.
Synthesis and fungicidal activities of new 1,2,4-triazolo[1,5-a]-
pyrimidines. Chem. Biodiversity 2009, 6, 1254−1265.
L
dx.doi.org/10.1021/jf503794g | J. Agric. Food Chem. XXXX, XXX, XXX−XXX

Journal of Agricultural and Food Chemistry
Article
(12) Jin, Y.-X.; Zhong, A.-G.; Zhang, Y.-J.; Pan, F.-Y. Synthesis,
crystal structure, spectroscopic properties, antibacterial activity and
theoretical studies of a novel difunctional acylhydrazone. J. Mol. Struct.
2011, 1002 (1−3), 45−50.
(13) Liu, X. H.; Liu, H. J.; Tan, C. X.; Weng, J. Q. Application of
acylhydrazonederivatives as fungicide. Faming Zhuanli Shenqing, CN
101874496 A 2010.
(14) Fuog, D.; Fergusson, S. J.; Fluckiger, C. Pymetrozine: A novel
̈
insecticide affecting aphids and whiteflies. In Insecticides with Novel
Modes of Action. Springer: Berlin Heidelberg, 1998; pp 40−49.
(15) Jose, L.; Armes, N. J.; Farlow, R.; Aldridge, K.; Robin, F.;
Tedeschi, L. Metaflumizone, a new broad-spectrum insecticide for
crop protection. Congress Proceedings 2007 of the XVI International
Plant Protection Congress; British Crop Protection Council: Hampshire,
U.K., 2007; Vol. 1, pp 74−81.
(16) Lym, R. G.; Christianson, K. M. Diflufenzopyr increases
perennial weed control with auxin herbicides. Proceedings of the Western
Society of Weed Science (USA); Western Society of Weed Science: Las
Cruces, NM, 1998.
(17) Yao, K.; Zhao, M.; Zhang, X. Y.; Wang, Y. J.; Li, L.; Zheng, M.
Q.; Peng, S. Q. A class of oral N-[(1S,3S)-1-methyl-1,2,3,4-tetrahydro-
β-carboline-3-carbonyl]-N′-(amino-acid-acyl)hydrazine: Discovery,
synthesis, in vitro anti-platelet aggregation/in vivo anti-thrombotic
evaluation and 3D QSAR analysis. Eur. J. Med. Chem. 2011, 46, 3237−
3249.
(18) Song, H. J.; Liu, Y. X.; Liu, Y. X.; Wang, L. Z.; Wang, Q. M.
Synthesis and antiviral and fungicidal activity evaluation of β-carboline,
dihydro-β-carboline, tetrahydro-β-carboline alkaloids, and their de-
rivatives. J. Agric. Food Chem. 2014, 62 (5), 1010−1018.
(19) Takemiya, A.; Hartwig, J. F. Palladium-Catalyzed Synthesis of
Aryl Ketones by Coupling of Aryl Bromides with an Acyl Anion
Equivalent. J. Am. Chem. Soc. 2006, 128 (46), 14800−14801.
(20) Joshi, S. D.; Vagdevi, H. M.; Vaidya, V. P.; Gadaginamath, G. S.
Synthesis of new 4-pyrrol-1-yl benzoic acid hydrazide analogs and
some derived oxadiazole, triazole and pyrrole ring systems: A novel
class of potential antibacterial and antitubercular agents. Eur. J. Med.
Chem. 2008, 43, 1989−1996.
(21) Wang, K. L.; Su, B.; Wang, Z. W.; Wu, M.; Li, Z.; Hu, Y. N.;
Fan, Z. J.; Mi, N.; Wang, Q. M. Synthesis and antiviral activities of
phenanthroindolizidine alkaloids and their derivatives. J. Agric. Food
Chem. 2010, 58 (5), 2703−2709.
(22) Zhao, H. P.; Liu, Y. X.; Cui, Z. P.; Beattie, D.; Gu, Y. C.; Wang,
Q. M. Design, Synthesis, and Biological Activities of Arylmethylamine
Substituted Chlorotriazine and Methylthiotriazine Compounds. J.
Agric. Food Chem. 2011, 59 (21), 11711−11717.
M
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