Synthesis of Novel Oxime Sulfonate Derivatives of 2′(2′,6′)-(Di)chloropicropodophyllotoxins as Insecticidal Agents

Article abstract of DOI:10.1021/acs.jafc.5b02036

To discover novel natural-product-based pesticidal agents, we prepared a series of oxime sulfonate derivatives of 2′(2′,6′)-(Di)chloropicropodophyllotoxins by structural modification of podophyllotoxin. Their structures were well-characterized by proton nuclear magnetic resonance (¹H NMR), high-resolution mass spectrometry (HRMS), optical rotation, and melting point. Moreover, the key steric structure of compound 5f was unambiguously determined by single-crystal X-ray diffraction. Additionally, their insecticidal activity was evaluated at 1 mg/mL against the pre-third-instar larvae of oriental armyworm (Mythimna separata Walker), a typical lepidopteran pest. Among all derivatives, compounds 4c, 5c, and 5d exhibited more promising insecticidal activity, with the final mortality rates greater than 60%, when compared to their precursor podophyllotoxin and the positive control, toosendanin. It demonstrated that introduction of the chlorine atom at the C-2′ or C-2′,6′ position on the E ring of picropodophyllotoxin or oxime sulfonate derivatives of picropodophyllotoxin was important for the insecticidal activity and introduction of a halogen (e.g., fluorine, chlorine, or bromine) atom-substituted phenylsulfonyl group on the oxime fragment of 2′(2′,6′)-(di)chloropicropodophyllones could lead to more promising compounds.

Full text of DOI:10.1021/acs.jafc.5b02036

Page 1 of 27
Journal of Agricultural and Food Chemistry
Synthesis of Novel Oxime Sulfonate Derivatives of
2′(2′,6′)ꢀ(di)Chloropicropodophyllotoxins as Insecticidal Agents
Rong Wang, Xiaoyan Zhi, Jie Li, and Hui Xu*
Research Institute of Pesticidal Design & Synthesis, College of Sciences, Northwest A&F
University, Yangling 712100, Shaanxi Province, People′s Republic of China.
* Author to whom correspondence should be addressed [(H.X.) telephone
+86(0)29ꢀ87091952; fax: +86(0)29ꢀ87091952; eꢀmail orgxuhui@nwsuaf.edu.cn].
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Abstract
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To discover novel naturalꢀproductꢀbased pesticidal agents, we prepared a series of oxime
sulfonate derivatives of 2′(2′,6′)ꢀ(di)chloropicropodophyllotoxins by structural modification
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of podophyllotoxin. Their structures were well characterized by H NMR, HRMS, optical
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rotation, and melting point. Moreover, the key steric structure of 5f was unambiguously
determined by singleꢀcrystal Xꢀray diffraction. Additionally, their insecticidal activity was
evaluated at 1 mg/mL against the preꢀthirdꢀinstar larvae of oriental armyworm (Mythimna
separata Walker), a typical lepidopteran pest. Among all derivatives, compounds 4c, 5c and
5d exhibited more promising insecticidal activity with the final mortality rates greater than
60%, when compared with their precursor podophyllotoxin and the positive control,
toosendanin. It demonstrated that introduction of the chlorine atom at the Cꢀ2′ or Cꢀ2′,6′
position on the Eꢀring of picropodophyllotoxin or oxime sulfonate derivatives of
picropodophyllotoxin was important for the insecticidal activity; and introduction of a
halogen (e.g., fluorine, chlorine, or bromine) atomꢀsubstituted phenylsulfonyl group on the
oxime fragment of 2′(2′,6′)ꢀ(di)chloropicropodophyllones could lead to more promising
compounds.
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KEYWORDS: podophyllotoxin, oxime sulfonate, insecticidal activity, naturalꢀproductꢀbased
insecticide, structural modification, Mythimna separata Walker
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INTRODUCTION
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Oriental armyworm, Mythimna separata Walker, is a typical lepidopteran pest. The
intermittent outbreaks of its larvae at very high densities can terribly result in complete crop
loss.1 For example, in 2012, the severe outbreak of 3rdꢀgeneration larvae of M. separata
widely occurred in China, and approximately 4 million hectares of crops (e.g., corn, rice,
wheat, etc.) were affected.2 Currently, chemical pesticides are still the effective method to
control insect pests in agriculture. However, increasing and longꢀterm application of synthetic
agrochemicals has resulted in the development of resistance in pest populations and
environmental problems.3ꢀ7 Therefore, search of the potential alternatives to control insect
pests has recently received much attention in the agricultural field.8ꢀ15
Podophyllotoxin (1, Figure 1), a naturally occurring aryltetralin cyclolignan, is isolated
from the roots and rhizomes of some Podophyllum and Juniperus species. Compound 1 has
been widely used as the lead molecule for chemical modifications to discover more potent
antitumor, insecticidal and antifungal agents.16 Additionally, Hu et al. found that once a
chlorine atom was introduced at the Cꢀ2' position of podophyllotoxin derivatives, the
corresponding compounds showed no significant cytotoxicity.17 More recently, we have
prepared a series of oxime sulfonate derivatives of picropodophyllotoxin18 (2, Figure 1) and
4α/βꢀacyloxyꢀ2′(2′,6′)ꢀ(di)halogenopodophyllotoxin derivatives19,20 (3, Figure 1) as
insecticidal agents, and found some derivatives exhibited more potent insecticidal activity
than toosendanin, a commercial botanical insecticide isolated from Melia azedarach. Based
upon the aboveꢀmentioned results, and in continuation of our program to discover new
potential naturalꢀproductꢀbased insecticidal agents, consequently, in the present paper we
prepared a series of novel oxime sulfonate derivatives of 2′ꢀchloropicropodophyllotoxin and
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2′,6′ꢀdichloropicropodophyllotoxin (4 and 5, Figure 1). Their insecticidal activity against M.
separata was also presented.
MATERIALS AND METHODS
Chemicals and Reagents. NꢀChlorosuccinimide (NCS), sodium hydride, hydroxylamine
hydrochloride, and sulfonyl chlorides were purchased from Aladdin Chemistry Co., Ltd.
(Shanghai, China). Chromium trioxide was purchased from Tianli Chemical Reagent Co., Ltd.
(Tianjin, China). N,NꢀDimethylformamide, ethyl acetate, petroleum ether, pyridine,
dichloromethane, absolute ethanol, tetrahydrofuran were analytical grade and purchased from
Bodi Chemical Co., Ltd. (Tianjin, China). Sodium carbonate (Na2CO3), anhydrous sodium
sulfate (Na2SO4), and sodium hydrogen sulfite (NaHSO3) were purchased from Kelong
Chemical Reagent Co., Ltd. (Chengdu, China). Sodium bicarbonate (NaHCO3) was
purchased from Guangdong Guanghua Chemical Factory Co., Ltd. (Shantou, China).
Podophyllotoxin was purchased from Gansu Gerui Medicinal Materials Co., Ltd. (Lanzhou,
China). Analytical thinꢀlayer chromatography (TLC) and preparative thinꢀlayer
chromatography (PTLC) were performed with silica gel plates using silica gel 60 GF254
(Qingdao Haiyang Chemical Co., Ltd., Qingdao, China). Silica gel column chromatography
was performed with silica gel 200ꢀ300 mesh (Qingdao Haiyang Chemical Co., Ltd., Qingdao,
China). Picropodophyllotoxin (6, 72% yield, Scheme 1) was prepared in the same way as in
our previous report.18
Instrumentation. Melting points (mp) were determined on a XTꢀ4 digital melting point
apparatus (Beijing Tech Instrument Co., Ltd., Beijing, China) and were uncorrected. Optical
rotation was measured on a Rudolph Research Analytical Autopol III automatic polarimeter.
Proton nuclear magnetic resonance spectra (1H NMR) were recorded in CDCl3 on a Bruker
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Avance III 500 MHz instrument using tetramethylsilane (TMS) as the internal standard.
Highꢀresolution mass spectra (HRꢀMS) were carried out with IonSpec 4.7 Tesla FTMS
instrument.
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General Procedure for Synthesis
of 2′ꢀChloropicropodophyllotoxin (7),
2′,6′ꢀDichloropicropodophyllotoxin (8). To a solution of 6 (2 mmol) in dry
N,Nꢀdimethylformamide (DMF, 5 mL) at 0 oC, a solution of Nꢀchlorosuccinimide (NCS, 1.15
or 2.0 equiv.) in dry DMF (4 mL) was added dropwise for 10 min. During one hour, the
solution was allowed to warm slowly from 0 oC to 28 oC. When the reaction was complete
checked by TLC analysis, the reaction mixture was diluted with water (15 mL), and extracted
with ethyl acetate (30 mL × 3). Subsequently, the combined organic phase was washed by
saturated aq. Na2CO3 (30 mL × 3) and brine (30 mL), dried over anhydrous Na2SO4,
concentrated in vacuo, and purified by silica gel column chromatography eluting with
petroleum ether/ethyl acetate (1:1, v/v) to afford 7 (747 mg, 90% yield) or 8 (705 mg, 85%
yield).
Data for 7: CAS: 1458601ꢀ21ꢀ2; Yield: 90%, white solid, m.p. 115ꢀ117 oC [lit. 116ꢀ117 oC]20;
[α] 20D = 5 (c 3.2 mg/mL, acetone); 1H NMR (500 MHz, CDCl3) δ: 7.09 (s, 1H, Hꢀ5), 6.61 (s,
1H, Hꢀ8), 6.18 (s, 1H, Hꢀ6′), 5.94 (s, 2H, OCH2O), 4.66 (d, J = 9.5 Hz, 1H, Hꢀ11), 4.47 (d, J
= 9.5 Hz, 1H, Hꢀ4), 4.40ꢀ4.43 (m, 1H, Hꢀ11), 4.38 (d, J = 5.0 Hz, 1H, Hꢀ1), 3.92 (s, 3H,
3′ꢀOCH3), 3.91(s, 3H, 5′ꢀOCH3), 3.80 (s, 3H, 4′ꢀOCH3), 3.27ꢀ3.30 (m, 1H, Hꢀ2), 2.90 (s, 1H,
4ꢀOH), 2.60ꢀ2.65 (m, 1H, Hꢀ3); HRMS (ESI): Calcd for C22H25ClNO8 ([M+NH4]+), 466.1263;
found, 466.1271.
Data for 8: CAS: 1458601ꢀ23ꢀ4; Yield: 85%, pale yellow solid, m.p. 155ꢀ157 oC [lit. 159ꢀ160
oC]20; [α] 20D = 29 (c 3.8 mg/mL, acetone); 1H NMR (500 MHz, CDCl3) δ: 7.15 (s, 1H, Hꢀ5),
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6.05 (s, 1H, Hꢀ8), 5.93 (s, 2H, OCH2O), 5.01 (d, J = 6.5 Hz, 1H, Hꢀ1), 4.77 (d, J = 10.0 Hz,
1H, Hꢀ4), 4.44ꢀ4.46 (m, 2H, Hꢀ11), 3.99 (s, 3H, 3′ꢀOCH3), 3.96 (s, 3H, 5′ꢀOCH3), 3.89 (s, 3H,
4′ꢀOCH3), 3.49 (dd, J = 9.0, 6.5 Hz, 1H, Hꢀ2), 2.59ꢀ2.64 (m, 1H, Hꢀ3); HRMS (ESI): Calcd
for C22H24Cl2NO8 ([M+NH4]+), 500.0873; found, 500.0876.
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General Procedure for Synthesis of 2′ꢀChloropicropodophyllone (9) and
2′,6′ꢀDichloropicropodophyllone (10). A mixture of 7 or 8 (1 mmol), chromium trioxide
(CrO3, 5 mmol), and pyridine (10 mmol) in dry dichloromethane (20 mL) was stirred at room
temperature. When the reaction was complete after 4 h, checked by TLC analysis, the mixture
was diluted by dichloromethane (60 mL), washed by saturated aq. NaHSO3 (30 mL) and
brine (30 mL), dried over anhydrous Na2SO4, concentrated under reduced pressure, and
purified by silica gel column chromatography eluting with petroleum ether/ethyl acetate (1:1,
v/v) to afford 9 (386 mg, 86% yield) or 10 (383 mg, 79% yield).
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Data for 9: Yield: 86%, yellow solid, m.p. 105ꢀ107 C; [α] 20D = ꢀ30 (c 3.4 mg/mL, acetone);
1H NMR (500 MHz, CDCl3) δ: 7.51 (s, 1H, Hꢀ5), 6.68 (s, 1H, Hꢀ8), 6.07 (d, J = 1.0 Hz, 2H,
OCH2O), 5.85 (s, 1H, Hꢀ6′), 5.21 (d, J = 1.5 Hz, 1H, Hꢀ1), 4.75 (d, J = 9.0 Hz, 1H, Hꢀ11),
4.32ꢀ4.35 (m, 1H, Hꢀ11), 3.95 (s, 3H, 3′ꢀOCH3), 3,85 (s, 3H, 5′ꢀOCH3), 3.54 (s, 3H, 4′ꢀOCH3),
3.42 (dd, J = 8.0, 2.0 Hz, 1H, Hꢀ2), 3.15ꢀ3.18 (m, 1H, Hꢀ3); HRMS (ESI): Calcd for
C22H20O8Cl ([M+H]+), 447.0841; Found, 447.0835.
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Data for 10: Yield: 79%, yellow solid, m.p. 90ꢀ92 C, [α] 20 = ꢀ67 (c 3.3 mg/mL, acetone);
D
1H NMR (500 MHz, CDCl3) δ: 7.48 (s, 1H, Hꢀ5), 6.32 (s, 1H, Hꢀ8), 6.01 (s, 2H, OCH2O),
5.68 (d, J = 4.5 Hz, 1H, Hꢀ1), 4.91 (dd, J = 9.0, 3.5 Hz, 1H, Hꢀ11), 4.50 (dd, J = 9.5, 7.0 Hz,
1H, Hꢀ11), 3.97 (s, 3H, 3′ꢀOCH3), 3.96 (s, 3H, 5′ꢀOCH3), 3.86 (s, 3H, 4′ꢀOCH3), 3.65ꢀ3.69 (m,
1H, Hꢀ2), 3.43ꢀ3.45 (m, 1H, Hꢀ3); HRMS (ESI): Calcd for C22H19O8Cl2 ([M+H]+), 481.0451;
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Found, 481.0455.
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General Procedure for Synthesis of Oximes of 2′ꢀChloropicropodophyllone and
2′,6′ꢀDichloropicropodophyllone (11 and 12). A mixture of 9 or 10 (0.5 mmol),
hydroxylamine hydrochloride (0.75 mmol), and pyridine (2 mmol) in absolute ethanol (20
mL) was refluxed. When the reaction was complete after 72 h, checked by TLC analysis, the
solvent was removed under reduced pressure and saturated aq. NaHCO3 (15 mL) was added
to the residue, which was extracted with ethyl acetate (3 × 30 mL). The combined organic
phase was dried over anhydrous Na2SO4, filtered, concentrated under reduced pressure, and
purified by silica gel column chromatography eluting with petroleum ether/ethyl acetate (1:1,
v/v) to afford 11 (193 mg, 83% yield) or 12 (214 mg, 89% yield).
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Data for 11：Yield: 83%, pale yellow solid, m.p. 106ꢀ107 C, [α] 20 = ꢀ27 (c 3.4 mg/mL,
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acetone); H NMR (500 MHz, CDCl3) δ: 7.27 (s, 1H, Hꢀ5), 6.70 (s, 1H, Hꢀ8), 5.99 (s, 2H,
OCH2O), 5.95 (s, 1H, Hꢀ6′), 5.07 (d, J = 2.0 Hz, 1H, Hꢀ1), 4.47ꢀ4.53 (m, 2H, Hꢀ11), 3.93 (s,
3H, 3′ꢀOCH3), 3.83ꢀ3.86 (m, 4H, Hꢀ3 and 5′ꢀOCH3), 3.52 (s, 3H, 4′ꢀOCH3), 3.43 (dd, J = 8.5,
2.5 Hz, 1H, Hꢀ2); HRMS (ESI): Calcd for C22H21O8NCl ([M+H]+), 462.0950; Found,
462.0949.
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Data for 12: Yield: 89%, white solid, m.p. 118ꢀ120 C, [α] 20 = 54 (c 5.3 mg/mL, acetone);
D
1H NMR (500 MHz, CDCl3) δ: 8.63 (s, 1H, OH), 7.41 (s, 1H, Hꢀ5), 6.21 (s, 1H, Hꢀ8), 5.93 (s,
2H, OCH2O), 5.24 (d, J = 10.5 Hz, 1H, Hꢀ1), 5.02ꢀ5.08 (m, 1H, Hꢀ11), 4.28ꢀ4.33 (m, 2H,
Hꢀ11, Hꢀ3), 3.99 (s, 3H, 4′ꢀOCH3), 3.93 (s, 3H, 3′ꢀOCH3), 3.90(s, 3H, 5′ꢀOCH3), 3.59 (dd, J =
10.5, 6.5 Hz, 1H, Hꢀ2); HRMS (ESI): Calcd for C22H20O8NCl2 ([M+H]+), 496.0560; Found,
496.0555.
General Procedure for Synthesis of Oxime Sulfonate Derivatives of
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2′(2′,6′)ꢀ(di)Chloropicropodophyllotoxins (4aꢀj, and 5c,d,fꢀk). To a stirred solution of NaH
(1.4 mmol) in dry tetrahydrofuran (8 mL) at −10 °C was slowly added compound 11 or 12
(0.2 mmol). After the completion of addition, the reaction mixture was stirred at −10 °C for
0.5 h. Then the corresponding sulfonyl chlorides RSO2Cl (13, 0.8 mmol) were added to the
above mixture. The reaction process was checked by TLC analysis. When the reaction
mixture was stirred for 3 h at −10 °C (for methanesulfonyl chloride, the reaction mixture was
stirred at room temperature for 48 h), saturated aq. NaHCO3 (15 mL) was added to the
mixture, which was extracted with dichloromethane (3×30 mL). The combined organic phase
was dried over anhydrous Na2SO4, filtered, concentrated under reduced pressure, and purified
by PTLC to give target products 4aꢀj, and 5c,d,fꢀk in 37−91% yields. The example data of
4aꢀe, and 5c,d are described as follows, whereas the data of 4fꢀj, and 5fꢀk are shown in the
Supporting Information.
Data for 4a: Yield: 82%, white solid, m.p. 102ꢀ104 oC, [α] 20D = 4 (c 4.2 mg/mL, acetone); 1H
NMR (500 MHz, CDCl3) δ: 8.02ꢀ8.04 (m, 2H, Hꢀ2", Hꢀ6"), 7.73 (t, J = 7.5 Hz, 1H, Hꢀ4"),
7.59ꢀ7.62 (m, 2H, Hꢀ3", Hꢀ ), 7.14 (s, 1H, Hꢀ5), 6.71 (s, 1H, Hꢀ8), 6.02 (s, 2H, OCH2O),
5"
5.77 (s, 1H, Hꢀ6'), 5.08 (d, J = 2.0 Hz, 1H, Hꢀ1), 4.54 (dd, J = 10.0, 7.0 Hz, 1H, Hꢀ11), 4.31
(d, J = 10.0 Hz, 1H, Hꢀ11), 3.92 (s, 3H, 3′ꢀOCH3), 3.89ꢀ3.91 (m, 1H, Hꢀ3), 3.84 (s, 3H,
5′ꢀOCH3), 3.40 (dd, J = 8.5, 2.5 Hz, 1H, Hꢀ2), 3.36 (s, 3H, 4′ꢀOCH3); HRMS (ESI): Calcd for
C28H25O10NClS ([M+H]+), 602.0882; Found, 602.0885.
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Data for 4b: Yield: 48%, yellow solid, m.p. 112ꢀ114 C, [α] 20 = 6 (c 4.0 mg/mL, acetone);
D
1H NMR (500 MHz, CDCl3) δ: 8.90 (s, 1H, Hꢀ2
7.5 Hz, 1H, Hꢀ4"), 7.87 (t, J = 7.5 Hz, 1H, Hꢀ ), 7.09 (s, 1H, Hꢀ5), 6.73 (s, 1H, Hꢀ8), 6.03 (s,
"), 8.59 (d, J = 7.5 Hz, 1H, Hꢀ6"), 8.36 (d, J =
5"
2H, OCH2O), 5.77 (s, 1H, Hꢀ6'), 5.09 (s, 1H, Hꢀ1), 4.54ꢀ4.57 (m, 1H, Hꢀ11), 4.31 (d, J = 10.0
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Hz, 1H, Hꢀ11), 3.93 (s, 3H, 3′ꢀOCH3), 3.89ꢀ3.90 (m, 1H, Hꢀ3), 3.85 (s, 3H, 5′ꢀOCH3),
3.42ꢀ3.46 (m, 4H, 4′ꢀOCH3, Hꢀ2); HRMS (ESI): Calcd for C28H24O12N2ClS ([M+H]+),
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647.07; Found, 647.07.
Data for 4c: Yield: 75%, yellow solid, m.p. 106ꢀ108 C, [α] 20 = 10 (c 3.6 mg/mL, acetone)
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1H NMR (500 MHz, CDCl3) δ: 8.04ꢀ8.07 (m, 2H, Hꢀ2", Hꢀ
6"), 7.27ꢀ7.30 (m, 2H, Hꢀ3", Hꢀ5"),
7.12 (s, 1H, Hꢀ5), 6.72 (s, 1H, Hꢀ8), 6.03 (s, 2H, OCH2O), 5.77 (s, 1H, Hꢀ6'), 5.08 (d, J = 2.0
Hz, 1H, Hꢀ1), 4.55 (dd, J = 10.0, 7.0 Hz, 1H, Hꢀ11), 4.32 (d, J = 9.5 Hz, 1H, Hꢀ11), 3.92 (s,
3H, 3′ꢀOCH3), 3.90 (m, 1H, Hꢀ3), 3.84 (s, 3H, 5′ꢀOCH3), 3.39ꢀ3.41 (m, 4H, 4′ꢀOCH3 and
Hꢀ2); HRMS (ESI): Calcd for C28H24O10NClFS ([M+H]+), 620.0788; Found, 620.0784.
Data for 4d: Yield: 82%, white solid, m.p. 88ꢀ90 oC, [α] 20D = 15 (c 4.0 mg/mL, acetone); 1H
NMR (500 MHz, CDCl3) δ: 7.96 (d, J = 7.0 Hz, 2H, Hꢀ2", Hꢀ6"), 7.59 (dd, J = 6.5, 2.0 Hz,
2H, Hꢀ3", Hꢀ5"), 7.13 (s, 1H, Hꢀ5), 6.72 (s, 1H, Hꢀ8), 6.03 (s, 2H, OCH2O), 5.75 (s, 1H, Hꢀ6'),
5.08 (d, J = 2.5 Hz, 1H, Hꢀ1), 4.55 (dd, J = 9.5, 7.0 Hz, 1H, Hꢀ11), 4.30ꢀ4.32 (m, 1H, Hꢀ11),
3.92 (s, 3H, 3′ꢀOCH3), 3.87ꢀ3.90 (m, 1H, Hꢀ3), 3.84 (s, 3H, 5′ꢀOCH3), 3.41 (dd, J = 8.5, 2.5
Hz, 1H, Hꢀ2), 3.38 (s, 3H, 4′ꢀOCH3); HRMS (ESI): Calcd for C28H24O10NCl2S ([M+H]+),
636.0492; Found, 636.0498.
Data for 4e: Yield: 57%, yellow solid, m.p. 116ꢀ118 oC, [α] 20D = 21 (c 3.0 mg/mL, acetone);
1H NMR (500 MHz, CDCl3) δ: 8.22ꢀ8.24 (m, 1H, Hꢀ6
"), 7.79ꢀ7.81 (m, 1H, Hꢀ3"), 7.52ꢀ7.59
(m, 2H, Hꢀ4", Hꢀ5"), 7.00 (s, 1H, Hꢀ5), 6.71 (s, 1H, Hꢀ8), 5.99 (d, J = 3.5 Hz, 2H, OCH2O),
5.76 (s, 1H, Hꢀ6'), 5.07 (d, J = 2.0 Hz, 1H, Hꢀ1), 4.52ꢀ4.61 (m, 2H, Hꢀ11), 3.94 (s, 3H,
3′ꢀOCH3), 3.89ꢀ3.92 (m, 1H, Hꢀ3), 3.87 (s, 3H, 5′ꢀOCH3), 3.44ꢀ3.47 (m, 1H, Hꢀ2), 3.40 (s,
3H, 4′ꢀOCH3); HRMS (ESI): Calcd for C28H24O10NBrClS ([M+H]+), 679.9987; Found,
679.9985.
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Data for 5c: Yield: 42%, yellow solid, m.p. 98ꢀ100 C, [α] 20 = 16 (c 3.0 mg/mL, acetone)
D
1H NMR (500 MHz, CDCl3) δ: 8.07ꢀ8.09 (m, 2H, Hꢀ2", Hꢀ
6"), 7.29ꢀ7.30 (m, 2H, Hꢀ3", Hꢀ5"),
7.27 (s, 1H, Hꢀ5), 6.22 (s, 1H, Hꢀ8), 5.99 (d, J = 10.0 Hz, 2H, OCH2O), 5.25 (d, J = 10.5 Hz,
1H, Hꢀ1), 4.92ꢀ4.97 (m, 1H, Hꢀ11), 4.21ꢀ4.27 (m, 2H, Hꢀ11, Hꢀ3), 3.97 (s, 3H, 4′ꢀOCH3),
3.91 (s, 3H, 3′ꢀOCH3), 3.87 (s, 3H, 5′ꢀOCH3), 3.48ꢀ3.52 (m, 1H, Hꢀ2); HRMS (ESI): Calcd
for C28H23O10NCl2FS ([M+H]+), 654.0398; Found, 654.0407.
Data for 5d: Yield: 44%, yellow solid, m.p. 96ꢀ98 oC, [α] 20D = 20 (c 3.8 mg/mL, acetone) 1H
NMR (500 MHz, CDCl3) δ: 7.98ꢀ8.00 (m, 2H, Hꢀ2", Hꢀ6"), 7.58ꢀ7.59 (m, 2H, Hꢀ3", Hꢀ5"),
7.30 (s, 1H, Hꢀ5), 6.22 (s, 1H, Hꢀ8), 5.99 (d, J = 9.5 Hz, 2H, OCH2O), 5.25 (d, J = 10.0 Hz,
1H, Hꢀ1), 4.91ꢀ4.97 (m, 1H, Hꢀ11), 4.24ꢀ4.27 (m, 2H, Hꢀ11, Hꢀ3), 3.97 (s, 3H, 4′ꢀOCH3),
3.92 (s, 3H, 3′ꢀOCH3), 3.87(s, 3H, 5′ꢀOCH3), 3.48ꢀ3.51 (m, 1H, Hꢀ2); HRMS (ESI): Calcd
for C28H23O10NCl3S ([M+H]+), 670.0103; Found, 670.0100.
Biological Assay. The insecticidal activity of compounds (1, 4aꢀj, 5c,d,fꢀk and 6ꢀ12) was
evaluated as the mortality rate values by leafꢀdipping method as described previously,18
against the preꢀthirdꢀinstar larvae of oriental armyworm (Mythimna separata Walker). For
each compound, 30 preꢀthirdꢀinstar larvae (10 larvae per group) were used. Acetone solutions
of compounds 1, 4aꢀj, 5c,d,fꢀk, 6ꢀ12 and toosendanin (a positive control) were prepared at 1
mg/mL. Fresh wheat leaf discs (1×1 cm) were dipped into the corresponding solution for 3 s,
then taken out and dried. Leaf discs treated with acetone alone were used as a blank control
group. Several pieces of treated leaf discs were kept in each dish (10 larvae were raised in
o
each dish), which was then placed in a conditioned room (25 ± 2 C, 65ꢀ80% relative
humidity (RH), 12 h/12 h (light/dark) photoperiod). If the treated leaf discs were consumed,
additional treated ones were added to the dish. After 48 h, compoundꢀsoaked leaves were
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removed, and the larvae were fed with untreated fresh wheat leaves thereafter until adult
emergence. The corrected mortality rate values were obtained by the formula
corrected mortality rate (%) = (T − C) × 100/ (100% − C)
Where T is the mortality rate in the tested compounds group, and C is the mortality rate in the
blank control group (T and C were expressed as percentages).
RESULTS AND DISCUSSION
Synthesis. As described in Scheme 1, 2′ꢀchloropicropodophyllotoxin (7) and
2′,6′ꢀdichloropicropodophyllotoxin (8) were smoothly prepared by reaction of 6 with the
different amount of Nꢀchlorosuccinimide (NCS). Further oxidation of the hydroxy group at
the Cꢀ4 position of 7 or 8 in the presence of CrO3 and pyridine gave
2′ꢀchloropicropodophyllone (9) and 2′,6′ꢀdichloropicropodophyllone (10), respectively.
Subsequently, oximes of 2′ꢀchloropicropodophyllone and 2′,6′ꢀdichloropicropodophyllone (11
and 12) were obtained by reaction of hydroxylamine hydrochloride with 9 and 10,
respectively. Finally, oxime sulfonate derivatives of 2′(2′,6′)ꢀ(di)chloropicropodophyllotoxins
(4aꢀj, and 5c,d,fꢀk) were afforded by reaction of sulfonyl chlorides with 11 and 12,
1
respectively. The structures of all target compounds were well characterized by H NMR,
HRMS, optical rotation, and melting point. Moreover, the singleꢀcrystal structure of 5f was
further confirmed by Xꢀray crystallography (Figure 2). It clearly demonstrated that the
substituents on the C=N double bond of 5f were present in trans configuration; two hydrogen
atoms at the Cꢀ2 and Cꢀ3 positions all adopted α configuration, that is, the configuration of
lactone (Dꢀring ) of 5f was cis; and two chlorine atoms were at the Cꢀ2’ and Cꢀ6’ positions of
5f. Crystallographic data (excluding structure factors) for the structure of 5f have been
deposited with the Cambridge Crystallographic Data Centre as supplementary publication
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number CCDC 1055791. Copies of the data can be obtained, free of charge, on application to
CCDC, 12 Union Road, Cambridge CB2 1EZ, UK [fax: +44 (0)1223 336033 or eꢀmail:
deposit@ccdc.cam.ac.uk].
Insecticidal Activity. As shown in Table 1, the insecticidal activity of 1, 4aꢀj, 5c,d,fꢀk and
6ꢀ12 against the preꢀthirdꢀinstar larvae of M. separata in vivo was evaluated as the mortality
rates at 1 mg/mL. Toosendanin was used as a positive control at 1 mg/mL, and leaves treated
with acetone alone were used as a blank control group. The corresponding mortality rates of
the treated groups after 35 days were higher than those after 5, 10, 15, 20 and 25 days as we
reported in our previous papers.18ꢀ20 As described in Figure 3, the lethal time for 50%
mortality of 8, 4cꢀg, and 5c,d,f,g were 32, 23, 26, 31, 29, 29, 24, 25, 27, and 28 days,
respectively. In addition, the symptoms of M. separata in the treated groups were
characterized as follows: some larvae with the slim and wrinkled bodies died at the larval
stage due to feeding too much treated leaves during the first 48 h (Figure 4); some larvae
molted to malformed pupae or died during the pupation period (Figure 5), and it was
noteworthy that more than half of the final mortality rates were generally occupied by the
mortality rates at this stage; malformed moths also appeared with imperfect wings (Figure 6).
It suggested that the podophyllotoxin derivatives possibly exhibited the antiꢀmolting hormone
effect. Among all derivatives, compounds 7, 8, 4cꢀg, and 5c,d,f,g exhibited insecticidal
activity equal to, or higher than, that of the positive control toosendanin. Especially 4c, 5c and
5d exhibited more promising insecticidal activity with the final mortality rates greater than
60%. For example, the final mortality rates of 4c, 5c and 5d were 62.1%, 65.5% and 62.1%,
respectively; whereas the final mortality rates of the precursor 1 and toosendanin were 37.9%
and 48.3%, respectively. Introduction of the chlorine atom at the Cꢀ2′ or Cꢀ2′,6′ position on
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the Eꢀring of 6 afforded the potent compounds 7 and 8 (48.3% for 7 and 51.7% for 8, and
34.5% for 6). Similarly, introduction of the chlorine atom at the Cꢀ2′ or Cꢀ2′,6′ position on
the Eꢀring of oxime sulfonate derivatives of picropodophyllotoxin (2) could also produce
more potent compounds as compared with 2.18 For example, the final mortality rates of
4a,b,d,g,i,j were 37.9%, 41.4%, 58.6%, 55.2%, 41.4% and 37.9%, respectively; whereas the
final mortality rates of the corresponding oxime sulfonate derivatives of
picropodophyllotoxin were 24.1%, 13.8%, 55.2%, 27.6%, 44.8% and 27.6%, respectively.18
To oxime sulfonate derivatives of 2′ꢀchloropicropodophyllotoxins (4aꢀj), introduction of a
halogen atom (such as a fluorine, chlorine, or bromine atom) on the phenylsulfonyl group of
4a resulted in more promising compounds 4cꢀg. The final mortality rates of 4cꢀg were 62.1%,
58.6%, 51.7%, 55.2%% and 55.2%, respectively; whereas the final mortality rate of 4a was
only
37.9%.
Interestingly,
to
oxime
sulfonate
derivatives
of
2′,6′ꢀdichloropicropodophyllotoxins (5c,d,fꢀk), introduction of a fluorine, chlorine, or
bromine atom on the phenylsulfonyl group could also give potent compounds 5c,d,f,g, the
final mortality rates of which were 65.5%, 62.1%, 58.6%, and 58.6%, respectively. On the
contrary, to 4aꢀj and 5c,d,fꢀk, whether introduction of nitro group (electronꢀwithdrawing
group) or methyl/ethyl group (electronꢀdonating group) on the phenylsulfonyl fragment all
afforded less potent compounds 4b, 4i, 4j, 5i and 5j. When the methylsulfonyl group was
introduced at the oxime fragment of 12, the corresponding compound 5k showed less potent
insecticidal activity as compared with 5c (44.8% for 5k and 65.5% for 5c).
In
conclusion,
a
series
of
oxime
sulfonate
derivatives
of
2′(2′,6′)ꢀ(di)chloropicropodophyllotoxins were prepared by structural modification of
podophyllotoxin. Their insecticidal activity was evaluated at 1 mg/mL against the
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296
297
298
preꢀthirdꢀinstar larvae of M. separata. The key steric structure of 5f was unambiguously
determined by singleꢀcrystal Xꢀray diffraction. Among all derivatives, compounds 4c, 5c and
5d exhibited more potent insecticidal activity with the final mortality rates greater than 60%.
It demonstrated that introduction of the chlorine atom at the Cꢀ2′ or Cꢀ2′,6′ position on the
Eꢀring of picropodophyllotoxin or oxime sulfonate derivatives of picropodophyllotoxin was
necessary for the insecticidal activity; and introduction of a halogen (e.g., fluorine, chlorine,
or bromine) atomꢀsubstituted phenylsulfonyl group on the oxime fragment of
2′(2′,6′)ꢀ(di)chloropicropodophyllones led to more promising compounds. It will pave the
way for further design and chemical modification of podophyllotoxin as botanical insecticidal
agents.
ASSOCIATED CONTENT
Supporting Information
Data on 1H NMR, HRMS, optical rotation, and melting point for the target compounds. The
Supporting Information is available free of charge on the ACS Publications website at DOI:
AUTHOR INFORMATION
Corresponding Author
*(H.X.) Phone/fax: +86(0)29ꢀ87091952. Eꢀmail: orgxuhui@nwsuaf.edu.cn.
Funding
The present research was supported by Special Funds of Central Colleges Basic Scientific
Research Operating Expenses (No.2452015096), and the National Natural Science
Foundation of China (No.31071737).
Notes
The authors declare no competing financial interest.
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REFERENCES
(1) Han, E.; Gatehouse, A. G. Genetics of precalling period in the oriental armyworm,
Mythimna separata (Walker) (Lepidoptera: Noctuidae), and implications for migration.
Evolution 1991, 45, 1502–1510.
(2) Zeng, J.; Jiang Y.; Liu, J. Analysis of the armyworm outbreak in 2012 and suggestions of
monitoring and forecasting. Plant Prot. 2013, 39, 117−121.
(3) Sun, J. Y.; Liang, P.; Gao, X. W. Crossꢀresistance patterns and fitness in
fufenozideꢀresistant diamondback moth, Plutella xylostella (Lepidoptera: Plutellidae).
Pest Manag. Sci. 2012, 68, 285−289.
(4) Saddiq, B.; Shad, S. A.; Khan, H. A. A.; Aslam, M.; Ejaz, M.; Afzal, M. B. S. Resistance
in the mealybug Phenacoccus solenopsis Tinsley (Homoptera: Pseudococcidae) in
Pakistan to selected organophosphate and pyrethroid insecticides. Crop Prot. 2014, 66,
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(5) Ferreira, C. B. S.; Andrade, F. H. N.; Rodrigues, A. R. S.; Siqueira, H. A. A.; Gondim, M.
G. C. Resistance in field populations of Tetranychus urticae to acaricides and
characterization of the inheritance of abamectin resistance. Crop Prot. 2015, 67, 77−83.
(6) Guillette Jr., L. J.; Iguchi, T. Life in a contaminated world. Science 2012, 337, 1614–1615.
(7) Xu, H. Comments to “Insecticide resistance after silent spring”;
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(8) Chu, S. S.; Jiang, G. H.; Liu, Z. L. Insecticidal compounds from the essential oil of
Chinese medicinal herb Atractylodes chinensis. Pest Manag. Sci. 2011, 67, 1253−1257.
(9) Cheng, S. S.; Lin, C. Y.; Chen, Y. J.; Chung, M. J.; Chang, S. T. Insecticidal activities of
Cunninghamia konishii Hayata against Formosan subterranean termite, Coptotermes
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formosanus (Isoptera: Rhinotermitidae). Pest Manag. Sci. 2014, 70, 1215−1219.
(10) Lin, L.; Mulholland, N.; Wu, Q.; Beattie, D.; Huang, S.; Irwin, D.; Clough, J.; Gu, Y.;
Yang, G. F. Synthesis and antifungal activity of novel sclerotiorin analogues. J. Agric.
Food Chem. 2012, 60, 4480−4491.
(10) Sousa, R. M. O. F.; Rosa, J. S.; Oliveira, L.; Cunha, A.; FernandesꢀFerreira, M.
Activities of Apiaceae essential oils against armyworm, Pseudaletia unipuncta
(Lepidoptera: Noctuidae). J. Agric. Food Chem. 2013, 61, 7661–7672.
(11) Diao, W. R.; Hu, Q. P.; Feng, S. S.; Li, W. Q.; Xu, J. G. Chemical composition and
antibacterial activity of the essential oil from Green Huajiao (Zanthoxylum schinifolium)
against selected foodborne pathogens. J. Agric. Food Chem. 2013, 61, 6044–6049.
(12) Zhang, Y.; Wang, J. S.; Wang, X. B.; Gu, Y. C.; Wei, D. D.; Guo, C.; Yang, M. H.;
Kong, L. Y. Limonoids from the fruits of Aphanamixis polystachya (Meliaceae) and
their biological activities. J. Agric. Food Chem. 2013, 61, 2171−2182.
(13) Wu, M.; Han, G. F.; Wang, Z. W.; Liu, Y. X.; Wang, Q. M. Synthesis and antiviral
activities of antofine analogues with different Cꢀ6 substituent groups. J. Agric. Food
Chem. 2013, 61, 1030–1035.
(14) Fan; L. L.; Guo, Y.; Zhi, X. Y.; Y, X.; Xu, H. Stereoselective synthesis of
2αꢀchloropicropodophyllotoxins and insecticidal activity of their esters against oriental
armyworm, Mythimna separata Walker. J. Agric. Food Chem. 2014, 62, 3726–3733.
(15) Seiber, J. N.; Coats, J.; Duke, S. O.; Gross, A. D. Biopesticides: state of the art and future
opportunities. J. Agric. Food Chem. 2014, 62, 11613–11619.
(16) Lv, M.; Xu, H. Recent advances in semisynthesis, biosynthesis, biological activities,
mode of action, and structureꢀactivity relationship of podophyllotoxins: an update
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(2008ꢀ2010). MiniꢀRev. Med. Chem. 2011, 11, 901−909.
(17) Hu, H.; Liu, S. Y.; Cheng, Y. C.; Lee, K. H.; Wang Z. Q. Antitumor agents. 123.
Synthesis and human DNA topoisomerase II inhibitory activity of 2'ꢀchloro derivatives
of etoposide and 4.beta.ꢀ(arylamino)ꢀ4'ꢀOꢀdemethylpodophyllotoxins. J. Med. Chem.
1992, 35, 866–871.
(18) Wang, Y.; Shao, Y. H.; Wang, Y. Y.; Fan, L. L.; Yu, X.; Zhi, X. Y.; Yang, C.; Qu, H.;Yao,
X. J.; Xu, H. Synthesis and quantitative structureꢀactivity relationship (QSAR) study of
novel isoxazoline and oxime derivatives of podophyllotoxin as insecticidal agents. J.
Agric. Food Chem. 2012, 60, 8435–8443.
(19) Che, Z. P.; Yu, X.; Zhi, X. Y.; Fan, L. L.; Xu, H. Synthesis of novel
4αꢀ(acyloxy)ꢀ2'(2',6')ꢀ(di)halogenopodophyllotoxin derivatives as insecticidal agents. J.
Agric. Food Chem. 2013, 61, 8148−8155.
(20) Che, Z. P.; Yu, X.; Fan, L. L.; Xu, H. Insight into dihalogenation of Eꢀring of
podophyllotoxins, and their acyloxyation derivatives at the C4 position as insecticidal
agents. Bioorg. Med. Chem. Lett. 2013, 23, 5592−5598.
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Figure Captions
Figure 1. Chemical structures of podophyllotoxin (1) and its derivatives (2ꢀ5).
Figure 2. Xꢀray crystal structure of 5f.
Figure 3. The lethal time for 50% mortality of 8, 4cꢀg, and 5c,d,f,g.
Figure 4. The representative abnormal larvae pictures of 4g (WRꢀ015), 4d (WRꢀ026), 4c
(WRꢀ039), 5g (WRꢀ022), 5j (WRꢀ043), 5d (WRꢀ038) and 5c (WRꢀ041) during the larval
period (CK: blank control group).
Figure 5. The representative malformed pupae pictures of 4g (WRꢀ015), 4d (WRꢀ026), 4f
(WRꢀ046), 5j (WRꢀ043), 5g (WRꢀ022), 5d (WRꢀ038) and 5c (WRꢀ041) during the pupation
period (CK: blank control group).
Figure 6. The representative malformed moth pictures of 4g (WRꢀ015), 4d (WRꢀ026), 4a
(WRꢀ030), 5k (WRꢀ036), 5d (WRꢀ038), 5c (WRꢀ041), and 4f (WRꢀ046) during the stage of
adult emergence (CK: blank control group).
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Figure 1.
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Figure 2.
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Figure 3.
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Figure 4.
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Figure 5.
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Figure 6.
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Journal of Agricultural and Food Chemistry
OH
N
O
O
O
O
O
NH2OH.HCl
EtOH/Py
O
O
13a-j
O
Cl
O
Cl
NaH/THF
-10 oC~rt
72 h, reflux
83%
O
O
O
O
O SO2R
O
O
9
N
48-91%
O
O
O
11
4 h, 20 oC
86%
CrO3/Py
CH2Cl2
O
Cl
OH
O
O
O
O
O
4a-j
O
O
Cl
NCS (1.15 equiv)
24 h, 0~28 oC
90%
OH
O
O
10% aq. AcONa
EtOH
O
O
O
O
1
18 h, 80 oC
O
7
OH
72%
NCS (2 equiv)
24 h, 0~28 oC
85%
O
O
O
O
O
O
6
O SO2R
O
Cl
N
Cl
O
O
O
O
O
O
8
O
13c,d,f-k
Cl
Cl
NaH/THF
4 h, 20 oC
79%
CrO3/Py
CH2Cl2
-10 oC~rt
O
O
OH
O
N
O
37-87%
O
5c,d,f-k
O
O
O
NH2OH.HCl
EtOH/Py
O
O
O
O
Cl
72 h, reflux
89%
Cl
Cl
Cl
O
O
O
O
O
12
O
10
R =
O
Br
Br
c
a
b
d
e
f
O
Cl
F
S
R
Cl
NO2
I
13a-k
g
i
CH3
k
h
j
Br
C2H5
H3C
Scheme
1.
Synthesis
of
oxime
sulfonate
derivatives
of
2′(2′,6′)ꢀ(di)chloropicropodophyllotoxins (4aꢀj, and 5c,d,fꢀk).
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Table
1.
Insecticidal
Activity
of
Oxime
Sulfonate
Derivatives
of
2′(2′,6′)ꢀ(di)Chloropicropodophyllotoxins (4aꢀj, and 5c,d,fꢀk) against M. separata on
Leaves Treated with a Concentration of 1 mg/mL.
corrected mortality rate (%)
compound
10 days
13.3 ± 3.3
10.0 ± 0
20 days
16.7 ± 3.3
16.7 ± 3.3
33.3 ± 3.3
30.0 ± 0
35 days
37.9 ± 0
1
6
34.5 ± 3.3
48.3 ± 0
7
16.7 ± 3.3
20.0 ± 0
8
51.7 ± 3.3
37.9 ± 5.8
34.5 ± 3.3
34.5 ± 3.3
31.0 ± 3.3
37.9 ± 5.8
41.4 ± 3.3
62.1 ± 3.3
58.6 ± 5.8
51.7 ± 3.3
55.2 ± 3.3
55.2 ± 3.3
44.8 ± 3.3
41.4 ± 3.3
37.9 ± 0
9
13.3 ± 3.3
10.0 ± 0
20.0 ± 0
10
16.7 ± 3.3
20.0 ± 0
11
13.3 ± 3.3
6.7 ± 3.3
3.3 ± 3.3
13.3 ± 3.3
16.7 ± 3.3
20.0 ± 0
12
13.3 ± 3.3
16.7 ± 3.3
26.7 ± 3.3
40.0 ± 0
4a
4b
4c
4d
33.3 ± 3.3
30.0 ± 0
4e
16.7 ± 3.3
20.0 ± 0
4f
26.7 ± 3.3
30.0 ± 0
4g
16.7 ± 3.3
16.7 ± 3.3
13.3 ± 3.3
6.7 ± 3.3
16.7 ± 3.3
16.7± 3.3
20.0 ± 0
4h
30.0 ± 0
4i
20.0 ± 0
4j
13.3 ± 3.3
46.7 ± 3.3
36.7 ± 3.3
33.3 ± 3.3
33.3 ± 3.3
23.3 ± 3.3
23.3 ± 3.3
23.3 ± 3.3
20.0 ± 0
5c
65.5 ± 3.3
62.1± 3.3
58.6 ± 5.8
58.6 ± 0
5d
5f
5g
16.7 ± 3.3
16.7 ± 3.3
16.7 ± 3.3
16.7 ± 3.3
10.0 ± 0
5h
44.8 ± 3.3
44.8 ± 3.3
41.4 ± 3.3
44.8 ± 3.3
48.3 ± 0
5i
5j
5k
toosendanin
blank control
13.3 ± 3.3
0 ± 0
20.0 ± 0
0 ± 0
3.3 ± 3.3
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