5-Methyl-1,2,3-thiadiazoles synthesized via ugi reaction and their fungicidal and antiviral activities

Article abstract of DOI:10.1021/jf1006193

1,2,3-Thiadiazoles, an important synthetic active substructure, are nowadays becoming one of the important branches in novel pesticide development. To develop pesticide candidates with diverse biological activities and probe their structure-activity relat

Full text of DOI:10.1021/jf1006193

7846 J. Agric. Food Chem. 2010, 58, 7846–7855
DOI:10.1021/jf1006193
5-Methyl-1,2,3-thiadiazoles Synthesized via Ugi Reaction
and Their Fungicidal and Antiviral Activities
Q
INGXIANG
Z
HENG, N
A
M
I
, ZHIJIN
F
AN,* XIANG
Z
UO, HAIKE
Z
HANG, HUAN
W
ANG
,
AND
Z
HIKUN
Y
ANG
State Key Laboratory of Elemento-Organic Chemistry, Nankai University, 94 Weijin Road, Nankai
District, Tianjin 300071, People’s Republic of China. These authors contributed equally to this work.
1,2,3-Thiadiazoles, an important synthetic active substructure, are nowadays becoming one of the
important branches in novel pesticide development. To develop pesticide candidates with diverse
biological activities and probe their structure-activity relationship, three series of 5-methyl-1,2,3-
thiadiazoles were rationally designed and synthesized using a simple and convenient one-step synthetic
procedure via Ugi reaction. Biological activities of the target compounds including fungicidal activity,
antivirus activity in vitro and in vivo, and systemic acquired resistance were systematically evaluated.
The results indicated that compound III10 showed broad-spectrum of activities against most fungi tested,
and compounds I10 and II17 showed excellent potential antivirus activities as compared to positive
control agent ribavirin. The preliminary structure-activity relationship was also discussed. The results of
these studies indicated that the 5-position-substituted 1,2,3-thiadiazoles exhibited good antivirus activity
and were worthy of further study in pesticide development.
KEYWORDS: 5-Methyl-1,2,3-thiadiazole; systemic acquired resistance; Ugi reaction; fungicide
INTRODUCTION
Part of our previous studies on lead derivation by Ugi four-
component condensation reaction (U-4CR) indicated that Ugi
four-component condensation is an efficient and convenient
green method to combine several active moieties into one mole-
cule, and it could be convenient to implement structural mod-
ification via alternation of diverse starting materials (12). On
the basis of the fact that 4-methyl-1,2,3-thiadiazole-containing
3-(trifluoromethyl)phenyl and 2-methylphenyl derivatives pos-
sessed potentially wide spectra of fungicidal activity, 4-methyl-
1,2,3-thiadiazole-containing 3-(trifluoromethyl)phenyl and
4-hydroxyphenyl derivatives possessed good potential direct anti-
virus activities against TMV in vitro and because replacement of
the Cl atom by a F atom improved their direct inhibition activi-
ties against TMV in vitro, 4-methyl-1,2,3-thiadiazole-containing
phenyl, 2-(trifluoromethyl)phenyl, 3-(trifluoromethyl)phenyl,
3-nitrophenyl, 4-nitrophenyl, 2-methylphenyl, and 4-hydroxy-
phenyl derivatives possessed good potential bioactivities in vivo
including protection, inactivation, curative, and induction acti-
vities against TMV (12), there was an expectation that the activity
of 5-methyl-1,2,3-thiadiazole also deserved further studies.
Moreover, the compounds obtained through U-4CR with
R-amino acylamide skeletons showed considerable bioactivities.
These are particularly useful with respect to the above parameters (iv).
Generally, U-4CR refers to the reaction among an amine, a
carbonyl compound (aldehyde or ketone), an isocyanide, and a
carboxylicacidtoprovide a singlecondensationproduct (14-17).
On the basis of an overall consideration of the above factors (i, ii,
and iii), 5-methyl-1,2,3-thiadiazole-4-carboxylic acid was synthe-
sized as the intermediate with carboxylic acid as the functional
group. To obtain compounds with potential bioactivities and
Derivatives of 1,2,3-thiadiazole with a wide variety of bio-
logical activities including antivirus activity (1, 2), systemic
acquired resistance (3, 4), fungicide activity (3, 5-8), insecticide
activity (9), and herbicide activity (10) were reported in the past
decade. Moreover, the properties of easy breakdown of the 1,2,3-
thiadiazole ring into low molecular weight compounds through
release of N2 favor the use of its derivatives as environmentally
friendly pesticide candidates with low toxicity (11). The chemistry
of 5-substituted 1,2,3-thiradiazoles has been well studied in our
previous work (3,12,13). These papers address the issues of these
compounds and their biological activities. The results proved that
our strategy for pesticide development by combining other
bioactive moieties with a 1,2,3-thiadiazole moiety at the 5-posi-
tion really did work. However, derivatives at the 4-position of the
1,2,3-thiadiazole ring were seldom synthesized and considered in
pesticide development.
On the basis of the consideration above, the following char-
acteristics are highly desirable for our studies on this field: (i) a
compound containing a 1,2,3-thiadiazole ring with a functional
group at the 4-position should be easily modified; (ii) as an
intermediate, this compound should be conveniently synthesized;
(iii) the substitution at the 4-position of the 1,2,3-thiadiazole ring
should be inert to most reactions so that the modifications at the
5-position would not be affected; (iv) a rapid and efficient
modification measure is preferred for our subsequent bioactivity
tests and structure-activity relationship studies.
*Author to whom correspondence should be addressed (phone
þ86-22-23504367; fax þ86-22-23503627; e-mail fanzj@nankai.edu.cn).
pubs.acs.org/JAFC
Published on Web 06/17/2010
© 2010 American Chemical Society

Article
J. Agric. Food Chem., Vol. 58, No. 13, 2010 7847
study the structure-activity relationship, like our previous
work (12), 3-chloro-4-methylbenzenamine and 3-fluoro-4-methyl-
benzenamine were chosen as the amine components, and sub-
stituted benzaldehydes were chosen as the carbonyl components;
two series including isopropyl isocyanide and cyclohexyl isocya-
nide were chosen as the isocyanide components, respectively. This
will provide useful information for the study of the structure-
activity relationship of these new structures.
To continue our ongoing program to develop 1,2,3-thiadia-
zoles with biologically interesting compounds, here we report the
synthesis and fungicidal activities, antivirus activity in vitro and in
vivo, and systemic acquired resistance of 5-methyl-1,2,3-thiadia-
zole-4-carboxamides including N-((isopropylcarbamoyl)(aryl)-
methyl)-N-(3-chloro-4-methylphenyl)-5-methyl-1,2,3-thiadiazole-
4-carboxamides (Series I), N-((isopropylcarbamoyl)(aryl)methyl)-
N-(3-fluoro-4-methylphenyl)-5-methyl-1,2,3-thiadiazole-4-carbox-
amides (Series II) and N-((cyclohexylcarbamoyl)(aryl)methyl)-
N-(3-fluoro-4-methylphenyl)-5-methyl-1,2,3-thiadiazole-4-carbox-
amides (Series III). The results indicated that compound III10
showed broad-spectrum of activities against several fungi tested,
and compound I10 and II17 showed excellent potential antivirus
activities as compared with ribavirin. The 5-position substituted
1,2,3-thiadiazoles are worthy of further studies. Their preliminary
structure-activity relationship is also discussed here.
as an eluent were performed to obtain the product. The yields, physical
properties, and 1H NMR and HRMS data of the target compounds are
listed in Table 1.
Biological Screening. Biological activities of the target compounds
including fungicide and direct antivirus activities such as protection,
inactivation, curative effect and systemic acquired resistance were eval-
uated systematically according to the standard operation practice (SOP) as
described in ref 3 and the following procedures.
Fungicide Screening. Preliminary screening was conducted by a fungal
growth inhibition method according to reference using potato dextrose
agar (PDA) as cultivation medium (3). Fungi used in this studies included
Alternaria solani (AS), Botrytis cinerea (BC), Cercospora arachidicola-
(CA), Cercospora beticola (CB), Colletotrichum lagenarium (CL), Fusar-
ium oxysporum (FO), Gibberella zeae (GZ), Puccinia triticina Eriks (PT),
Phytophthora infestans (Mont) de Bary (PI), Physalospora piricola (PP),
Pellicularia sasakii (PS), and Rhizoctonia solani (RS).
Systemic Acquired Resistance Screening. Systemic acquired resistance
of the target compounds was detected using tobacco against the tobacco
mosaic virus (TMV) system as described in ref 3. The induction activity
was evaluated using the antivirus inhibition ratio, which was calculated by
the average number of viral inflammations on the inoculated leaves with
the corresponding control accordingly. Tiadinil (TDL) and ribavirin were
chosen as positive control and negative control, respectively, and all
compounds tested were conducted at 100 and 50 μg/mL, respectively.
Protective Effects of the Target Compounds against TMV in Vivo.
Healthy fresh tobacco leaves growing at six-leaf age were selected for the
tests. The compound solution was smeared on the whole leaves, and then
the leaves were dried in the greenhouse. After 12 h, TMV at a concentra-
tion of 5.88 ꢀ 10-2 μg/mL was inoculated on the upper three leaves using
the conventional juice-robbing method, and the solvent was smeared on
the lower three leaves as a control. The local lesion numbers were then
recorded 2-3 days after inoculation. For each compound, three repeti-
tions were conducted. Most compounds tested were conducted at 500 and
100 μg/mL, respectively. Only a few compound tests were conducted at
500 and 200 μg/mL, respectively.
Inactivation Effect of the Target Compounds against TMV in Vivo.
Healthy fresh tobacco leaves growing at six-leaf age were selected for the
tests. TMV virus at a concentration of 5.88 ꢀ 10-2 μg/mL was inhibited by
mixing with the target compound solution at the same volume for 30 min.
Then the mixture was inoculated on the upper three leaves using the
conventional juice-robbing method, and the solvent was smeared on the
lower three leaves as a control. The local lesion numbers were then
recorded 2-3 days after inoculation. For each compound, three repeti-
tions were conducted. All compounds tested were conducted at 500 and
100 μg/mL, respectively.
EXPERIMENTAL METHODS
Equipment for Structural Characterization. Melting points of all
compounds were determined on an X-4 binocular microscope (Gongyi
Tech. Instrument Co., Henan, China), and the thermometer was not
corrected. Proton NMR spectra were obtained using a Bruker Avance-400
MHz spectrometer, and chemical shift values (δ) werereported as parts per
million with deutero-dimethyl sulfoxide (DMSO-d6) or CDCl3 as the
solvent and tetramethylsilane (TMS) as the internal standard. HRMS data
were obtained on an FTICR-MS Varian 7.0T FTICR-MS instrument. All
solvents and liquid reagents were of analytical reagent grade and were
dried in advance and distilled before use. Flash column chromatography
purification was carried out by using silica gel.
Preparation of Isopropyl Isocyanide and Cyclohexyl Isocyanide.
N-Isopropylformamide was prepared by reflux of 78.2 g (1.3 mol) of
isopropylamine and 130.0 g (1.8 mol) of ethyl formate for 16 h; 91.1 g of
N-isopropylformamide was collected through distillation under reduced
pressure with a yield of 79%. A solution of 43.6 g (0.5 mol) of iso-
propylformamide in 237.3 g (3.0 mol) of refined pyridine was cooled to
0 °C, followed by the addition of 46.1 g (0.3 mol) of phosphorus oxychlo-
ride dropwise for 1 h under an ice bath. After the solution was then heated
to 60 °C and stirred for 30 min, it was cooled to 0-5 °C again. Fifty
milliliters of pentane and 200 mL of ice-water were added to the solution
with violent stirring. The organic phase was collected, and the aqueous
phase was extracted with 30 mL portions of pentane by four times. The
organic phase was dried by sodium sulfate, and 10.88 g of colorless foul-
smelling liquid isopropylisonitrile was obtained by distillation with a yield
of 31%. 1H NMR (CDCl3, 400 MHz): δ 1.37-1.39 (m, 6H, J = 2.0 Hz,
CH3), 3.72-3.80 (m, 1H, CH).
The preparation of cyclohexyl isocyanide was conducted according to
the procedures as described in refs 12 and 18. Cyclohexylisocyanide was
obtained as a colorless foul-smelling liquid with a yield of 73%.
Preparation of 5-Methyl-1,2,3-thiadiazolyl-4-carboxylic Acid.
5-Methyl-1,2,3-thiadiazole-4-carboxylic acid was prepared according to
the description in ref 19.
General Synthetic Procedure for 5-Methyl-1,2,3-thiadiazole-
4-carboxamides via U-4CR. 3-Chloro-4-methylbenzenamine (0.47 g,
3.0 mmol), 5-methyl-1,2,3-thiadiazole-4-carboxylic acid (0.43 g, 3.0 mmol),
and isopropyl isocyanide (0.20 g, 2.8 mmol) were added to a solution of
benzaldehyde (0.35 g, 3.0 mmol) in 10 mL of methanol in sequence. The
color of the mixture grew darker during this procedure. The solution was
then stirred for another 1 h at room temperature; afterward, removal of
solvent under reduced pressure and purification by flash column chromato-
graphy onsilicagel using ethyl acetate and petroleum ether (60-90 °C; 1:3)
Curative Effect of Target Compounds on TMV in Vivo. Healthy fresh
tobacco leaves growing at six-leaf age were selected for the tests. TMV at a
concentration of 5.88 ꢀ 10-2 μg/mL was inoculated on the whole leaves
using the conventional juice-robbing method. After the leaves were dried
in the greenhouse, the compound solution was smeared on the upper three
leaves, and the solvent was smeared on the lower three leaves as control.
The local lesion numbers were then recorded 2-3 days after inoculation.
For each compound, three repetitions were conducted. All compounds
tested were conducted at 500 and 100 μg/mL, respectively.
The activities of protection, inactivation, and curative effects against
TMV were calculated by the average number of viral inflammations on the
inoculated leaves with the corresponding control according to eq 1
CK - A
CK
Y ¼
ꢀ 100
ð1Þ
where Y is the antivirus inhibition ratio (protection, inactivation, and
curative effects in vivo) (%), CK is the average number of viral inflamma-
tions on the control leaves in vivo, and A is the average number of viral
inflammations on the target compound treated leaves in vivo.
RESULTS AND DISCUSSION
Preparation of the New Derivatives of 5-Methyl-1,2,3-thiadia-
zole-4-carboxylic Acid via U-4CR. All structures of the title com-
1
pounds synthesized were confirmed by H NMR and HRMS.

7848 J. Agric. Food Chem., Vol. 58, No. 13, 2010
Zheng et al.
Table 1. Yields, Physical Properties, and 1H NMR and HRMS Data of the Target Compounds
compd yield (%) appearance and mp (°C)
1H NMR, δ (solvent)
HRMS (m/z)
I1
71
white crystal, 163
1.15 (dd, 6H, J = 6.6 Hz, J = 24.6 Hz, CH3), 2.15 (s, 3H, Ph-CH3), 2.70 (s, 3H, CH3),
4.13-4.21 (m, 1H, J = 6.8 Hz, CH), 5.51 (d, 1H, J = 7.6 Hz, NH), 6.20 (s, 1H, PhCH),
6.81-7.28 (m, 7H, Ph) (CDCl3)
1.19 (dd, 6H, J = 6.8 Hz, J = 29.6 Hz, CH3), 2.18 (s, 3H, Ph-CH3), 2.70 (s, 3H, CH3),
4.14-4.22 (m, 1H, J = 6.8 Hz, CH), 5.86 (d, 1H, J = 7.6 Hz, NH), 6.23 (s, 1H, PhCH),
6.80-7.82 (m, 3H, Ph-H), 7.49 (d, 2H, J = 8.8 Hz, Ph-H), 8.12 (d, 2H, J = 8.8 Hz,
Ph-H) (CDCl3)
1.204 (dd, 6H, J = 6.8 Hz, J = 29.6 Hz, CH3), 2.177 (s, 3H, Ph-CH3), 2.722 (s, 3H, CH3),
4.152-4.237 (m, 1H, J = 6.8 Hz, CH), 5.872 (d, 1H, J = 7.6 Hz, NH), 6.261 (s, 1H, PhCH),
6.839-7.129 (m, 3H, Ph-H), 7.457 (t, 1H, J = 8.0 Hz, Ph-H), 7.633 (d, 1H, J = 7.6 Hz, Ph-H),
8.138-8.154 (d, 2H, Ph-H) (CDCl3)
[M þ Naþ]: 465.1128;
found: 465.1119
I2
36
light yellow crystal,
203-204
[M þ Naþ]: 510.0979;
found: 510.0973
I3
68
light yellow crystal,
189-190
[M þ Naþ]: 510.0979;
found: 510.0970
I4
72
52
59
52
65
53
61
41
white crystal, 178-179 1.15 (dd, 6H, J = 6.8 Hz, J = 24.6 Hz, CH3), 2.15 (s, 3H, Ph-CH3), 2.73 (s, 3H, CH3),
4.09-4.19 (m, 1H, J = 6.8 Hz, CH), 5.89 (d, 1H, J = 7.6 Hz, NH), 6.77 (s, 1H, PhCH),
6.86-7.95 (m, 7H, Ph-H) (CDCl3)
white crystal, 170-171 1.16 (dd, 6H, J = 6.6 Hz, J = 27.4 Hz, CH3), 2.18 (s, 3H, Ph-CH3), 2.69 (s, 3H, CH3),
4.11-4.21 (m, 1H, J = 6.8 Hz, CH), 5.59 (d, 1H, J = 7.6 Hz, NH), 6.14 (s, 1H, PhCH),
6.80-7.26 (m, 7H, Ph-H) (CDCl3)
white crystal, 165-166 1.17 (dd, 6H, J = 6.6 Hz, J = 30.0 Hz, CH3), 2.18 (s, 3H, Ph-CH3), 2.71 (s, 3H, CH3),
4.13-4.23 (m, 1H, J = 6.8 Hz, CH), 5.61 (d, 1H, J = 7.6 Hz, NH), 6.12 (s, 1H, PhCH),
6.83-7.27 (m, 7H, Ph-H) (CDCl3)
white crystal, 162-163 1.19 (dd, 6H, J = 6.4 Hz, J = 35.6 Hz, CH3), 2.11 (s, 3H, Ph-CH3), 2.68 (s, 3H, CH3),
4.16-4.24 (m, 1H, J = 6.8 Hz, CH), 5.65 (br, 1H, J = 7.6 Hz, NH), 6.63 (s, 1H, PhCH),
6.76-7.36 (m, 7H, Ph-H) (CDCl3)
[M þ Naþ]: 510.0979;
found: 510.0972
I5
[M þ Naþ]: 499.0738;
found: 499.0726
I6
[M þ Naþ]: 499.0738;
found: 499.0735
I7
[M þ Naþ]: 499.0738;
found: 499.0736
I8
white crystal, 172-173 1.15 (dd, 6H, J = 6.8 Hz, J = 29.6 Hz, CH3), 2.16 (s, 3H, Ph-CH3), 2.69 (s, 3H, CH3),
4.11-4.19 (m, 1H, J = 6.8 Hz, CH), 5.61 (d, 1H, J = 7.6 Hz, NH), 6.19 (s, 1H, PhCH),
6.79-7.22 (m, 7H, Ph-H) (CDCl3)
[M þ Naþ]: 483.1034;
found: 483.1024
I9
white crystal, 164
1.17 (dd, 6H, J = 6.4 Hz, J = 24.8 Hz, CH3), 2.17 (s, 3H, Ph-CH3), 2.70 (s, 3H, CH3),
4.11-4.21 (m, 1H, J = 6.8 Hz, CH), 5.64 (d, 1H, J = 7.6 Hz, NH), 6.11 (s, 1H, PhCH),
6.85-7.30 (m, 7H, Ph-H) (CDCl3)
1.19 (dd, 6H, J = 6.4 Hz, J = 29.2 Hz, CH3), 2.14 (s, 3H, Ph-CH3), 2.70 (s, 3H, CH3),
4.15-4.24 (m, 1H, J = 6.8 Hz, CH), 5.67 (br, 1H, NH), 6.48 (s, 1H, PhCH),
6.81-7.32 (m, 7H, Ph-H) (CDCl3)
[M þ Naþ]: 483.1034;
found: 483.1025
I10
white needle crystal,
167-168
[M þ Naþ]: 483.1034;
found: 483.1030
I11
white needle crystal, 203 1.17 (dd, 6H, J = 6.4 Hz, J = 27.2 Hz, CH3), 2.18 (s, 3H, Ph-CH3), 2.70 (s, 3H, CH3),
4.10-4.22 (m, 1H, J = 6.8 Hz, CH), 5.69 (d, 1H, J = 7.2 Hz, NH), 6.19 (s, 1H, PhCH),
6.81-7.32, 7.80-7.82 (m, 3H, Ph-H), 7.41 (d, 2H, J = 8.4 Hz, Ph-H),
7.54 (d, 2H, J = 8.4 Hz, Ph-H) (CDCl3)
white needle crystal,
170-171
[M þ Naþ]: 533.1002;
found: 533.1004
I12
63
51
1.18 (dd, 6H, J = 6.8 Hz, J = 33.2 Hz, CH3), 2.16 (s, 3H, Ph-CH3), 2.71 (s, 3H, CH3),
4.14-4.23 (m, 1H, J = 6.8 Hz, CH), 5.68 (d, 1H, J = 7.2 Hz, NH),
6.26 (s, 1H, PhCH), 6.81-7.55 (m, 7H, Ph-H) (CDCl3)
1.15 (dd, 6H, J = 6.8 Hz, J = 58.8 Hz, CH3), 2.14 (s, 3H, Ph-CH3),
2.71 (s, 3H, CH3), 4.11-4.21 (m, 1H, J = 6.8 Hz, CH), 5.36 (d, 1H,
J = 7.6 Hz, NH), 6.61 (s, 1H, PhCH), 6.78-7.17 (m, 3H, Ph-H),
7.16 (d, 1H, J = 8.0 Hz, Ph-H), 7.25 (t, 1H, J = 7.6 Hz, Ph-H),
7.37 (t, 1H, J = 7.6 Hz, Ph-H), 7.70 (d, 1H, J = 8.0 Hz, Ph-H) (CDCl3)
[M þ Naþ]: 533.1002;
found: 533.0996
I13
white solid, 113-115
[M þ Naþ]: 533.1002;
found: 533.0988
I14
52
white crystal, 145-146 1.14 (dd, 6H, J = 6.6 Hz, J = 27.0 Hz, CH3), 2.17 (s, 3H, Ph-CH3),
2.31 (s, 3H, Ph-CH3), 2.70 (s, 3H, CH3), 4.12-4.22 (m, 1H, J = 6.8 Hz, CH),
5.47 (d, 1H, J = 7.6 Hz, NH), 6.16 (s, 1H, PhCH), 6.82-7.16
[M þ Naþ]: 479.1284;
found: 479.1275
(m, 7H, Ph-H) (CDCl3)
I15
I16
I17
46
27
33
white crystal, 148-149 1.14 (dd, 6H, J = 6.4 Hz, J = 23.6 Hz, CH3), 2.16 (s, 3H, Ph-CH3), 2.27 (s, 3H, Ph-CH3),
2.70 (s, 3H, CH3), 4.12-4.21 (m, 1H, J = 6.8 Hz, CH), 5.51 (br, 1H, NH),
6.16 (s, 1H, PhCH), 6.81-7.15 (m, 7H, Ph-H) (CDCl3)
white crystal, 189-190 1.16 (dd, 6H, J = 6.8 Hz, J = 24.8 Hz, CH3), 2.11 (s, 3H, Ph-CH3),
2.49 (s, 3H, Ph-CH3), 2.69 (s, 3H, CH3), 4.15-4.24 (m, 1H, J = 6.8 Hz, CH),
5.41 (d, 1H, J = 7.2 Hz, NH), 6.57 (s, 1H, PhCH), 6.73-7.19 (m, 7H, Ph-H) (CDCl3)
white crystal, 178-179 1.15 (dd, 6H, J = 6.8 Hz, J = 27.2 Hz, CH3), 2.16 (s, 3H, Ph-CH3), 2.70 (s, 3H, CH3),
4.11-4.21 (m, 1H, J = 6.8 Hz, CH), 5.53 (d, 1H, J = 7.6 Hz, NH), 5.75 (s, 1H, OH),
6.13 (s, 1H, PhCH), 6.72 (d, 2H, J = 8.4 Hz, Ph-H), 6.84-6.80 (m, 2H, Ph-H),
7.07 (d, 2H, J = 8.4 Hz, Ph-H), 7.16 (br, 1H, Ph-H) (CDCl3)
white crystal, 203-204 1.13 (dd, 6H, J = 6.4 Hz, J = 23.2 Hz, CH3), 2.16 (s, 3H, Ph-CH3), 2.67 (s, 3H, CH3),
4.11-4.22 (m, 1H, J = 6.8 Hz, CH), 5.86 (d, 1H, J = 8.0 Hz, NH), 6.10 (s, 1H, PhCH),
6.64-6.84 (m, 5H, Ph-H), 6.94 (s, 1H, OH), 7.09 (t, 1H, J = 7.8 Hz, Ph-H),
7.209 (s, 1H, Ph-H) (CDCl3)
white crystal, 178-179 1.22 (dd, 6H, J = 6.4 Hz, J = 15.2 Hz, CH3), 2.19 (s, 3H, Ph-CH3), 2.71 (s, 3H, CH3),
4.15-4.23 (m, 1H, J = 6.8 Hz, CH), 6.19 (s, 1H, PhCH), 6.21 (s, 1H, OH),
6.76-7.21 (m, 7H, Ph-H) (CDCl3)
[M þ Naþ]: 479.1284;
found: 479.1276
[M þ Hþ]: 457.1465;
found: 457.1454
[M þ Naþ]: 481.1077;
found: 481.1070
I18
51
[M þ Naþ]: 481.1077;
found: 481.1072
I19
19
52
[M þ Naþ]: 481.1077;
found: 481.1078
II1
white crystal, 172-173 1.16 (dd, 6H, J = 6.8 Hz, J = 24.0 Hz, CH3), 2.06 (d, 3H, J = 1.6 Hz, Ph-CH3),
2.71 (s, 3H, CH3), 4.13-4.23 (m, 1H, CH), 5.49 (d, 1H, J = 7.6 Hz, NH),
6.23 (s, 1H, Ph-CH), 6.72-6.87 (m, 3H, Ph-H), 7.21-7.28 (m, 5H, Ph-H) (CDCl3)
[M þ Naþ]: 449.1423;
found: 449.1423

Article
J. Agric. Food Chem., Vol. 58, No. 13, 2010 7849
1H NMR, δ (solvent)
HRMS (m/z)
Table 1. Continued
compd yield (%) appearance and mp (°C)
II2
59
white crystal, 174
1.20 (dd, 6H, J = 6.4 Hz, J = 30.0 Hz, CH3), 2.08 (s, 3H, Ph-CH3), 2.71
(s, 3H, CH3), 4.13-4.23 (m, 1H, CH), 5.80 (d, 1H, J = 7.6 Hz, NH),
6.26 (s, 1H, Ph-CH), 6.69-6.90 (m, 3H, Ph-H), 7.49 (d, 2H, J = 8.4 Hz,
Ph-H), 8.12 (d, 2H, J = 8.8 Hz, Ph-H) (CDCl3)
[M þ Naþ]: 494.1274;
found: 494.1261
II3
46
white crystal, 156-157 1.19 (dd, 6H, J = 6.4 Hz, J = 30.4 Hz, CH3), 2.06 (s, 3H, Ph-CH3),
2.71 (s, 3H, CH3), 4.15-4.23 (m, 1H, CH), 5.92 (d, 1H,
[M þ Naþ]: 494.1274;
found: 494.1266
J = 7.6 Hz, NH), 6.31 (s, 1H, Ph-CH), 6.73-6.84
(m, 3H, Ph-H), 7.44 (t, 1H, J = 8.0 Hz, Ph-H), 7.62 (d, 1H, J = 7.6 Hz, Ph-H),
8.12-8.14 (d, 2H, Ph-H) (CDCl3)
1.16 (dd, 6H, J = 6.4 Hz, J = 24.8 Hz, CH3), 2.05 (s, 3H, Ph-CH3), 2.75 (s, 3H, CH3),
4.13-4.21 (m, 1H, CH), 5.79 (d, 1H, J = 7.2 Hz, NH), 6.78-6.91
(m, 4H, Ph-CH and Ph), 7.47 (t, 1H, J = 8.0 Hz, Ph-H), 7.43-7.96
(m, 4H, Ph-H) (CDCl3)
1.16 (dd, 6H, J = 6.4 Hz, J = 26.8 Hz, CH3), 2.08 (d, 3H, J = 1.6 Hz, Ph-CH3), 2.70
(s, 3H, CH3), 4.14-4.23 (m, 1H, CH), 5.51 (d, 1H, J = 7.6 Hz, NH), 6.17 (s, 1H, Ph-CH),
6.70-6.89 (m, 3H, Ph-H), 7.17-7.25 (m, 4H, Ph-H) (CDCl3)
1.18 (dd, 6H, J = 6.4 Hz, J = 25.6 Hz, CH3), 2.08 (d, 3H, J = 1.6 Hz, Ph-CH3), 2.71
(s, 3H, CH3), 4.13-4.23 (m, 1H, CH), 5.57 (d, 1H, J = 8.0 Hz, NH), 6.14 (s, 1H, Ph-CH),
6.73-6.88 (m, 3H, Ph-H), 7.11-7.28 (m, 4H, Ph-H) (CDCl3)
1.19 (dd, 6H, J = 6.4 Hz, J = 35.2 Hz, CH3), 2.02 (s, 3H, Ph-CH3), 2.68 (s, 3H, CH3),
4.16-4.25 (m, 1H, CH), 5.62 (d, 1H, J = 8.0 Hz, NH), 6.65 (s, 1H, Ph-CH),
6.70-6.95 (m, 3H, Ph-H), 7.03-7.36 (m, 4H, Ph-H) (CDCl3)
1.16 (dd, 6H, J = 6.4 Hz, J = 28.4 Hz, CH3), 2.08 (s, 3H, Ph-CH3), 2.70
(s, 3H, CH3), 4.16-4.23 (m, 1H, CH), 5.52 (d, 1H, J = 6.8 Hz, NH),
6.21 (s, 1H, Ph-CH), 6.69-6.85 (m, 3H, Ph-H), 6.92-7.23
(m, 4H, Ph-H) (CDCl3)
1.17 (dd, 6H, J = 6.4 Hz, J = 25.2 Hz, CH3), 2.07 (s, 3H, Ph-CH3),
2.70 (s, 3H, CH3), 4.12-4.21 (m, 1H, CH), 5.66 (d, 1H, J = 7.6 Hz, NH),
6.17 (s, 1H, Ph-CH), 6.73-7.25 (m, 7H, Ph-H) (CDCl3)
1.18 (dd, 6H, J = 6.4 Hz, J = 29.2 Hz, CH3), 2.04 (d, 3H, J = 0.8 Hz, Ph-CH3),
2.69 (s, 3H, CH3), 4.16-4.24 (m, 1H, CH), 5.67 (d, 1H, J = 7.2 Hz, NH),
6.50 (s, 1H, Ph-CH), 6.76-7.24 (m, 7H, Ph-H) (CDCl3)
1.18 (dd, 6H, J = 6.4 Hz, J = 26.0 Hz, CH3), 2.08 (s, 3H, Ph-CH3), 2.71
(s, 3H, CH3), 4.14-4.22 (m, 1H, CH), 5.65 (d, 1H, J = 7.2 Hz, NH), 6.21
(s, 1H, Ph-CH), 6.70-6.90 (m, 3H, Ph-H), 7.41 (d, 2H, J = 8.0 Hz, Ph-H),
7.54 (d, 2H, J = 8.0 Hz, Ph-H) (CDCl3)
1.18 (dd, 6H, J = 6.8 Hz, J = 32.4 Hz, CH3), 2.07 (s, 3H, Ph-CH3), 2.72 (s, 3H, CH3),
4.15-4.23 (m, 1H, CH), 5.63 (d, 1H, J = 8.0 Hz, NH), 6.28 (s, 1H, Ph-CH), 6.69-6.82
(m, 3H, Ph-H), 7.37-7.55 (m, 4H, Ph-H) (CDCl3)
1.15 (dd, 6H, J = 6.8 Hz, J = 57.2 Hz, CH3), 2.04 (s, 3H, Ph-CH3), 2.71 (s, 3H, CH3),
4.12-4.20 (m, 1H, CH), 5.35 (d, 1H, J = 7.2 Hz, NH), 6.62 (s, 1H, Ph-CH), 6.74
(s, 3H, Ph-H), 7.16-7.25 (m, 2H, Ph-H), 7.37 (t, 1H, J = 7.2 Hz, Ph-H), 7.70
(d, 1H, J = 8.0 Hz, Ph-H) (CDCl3)
1.14 (dd, 6H, J = 6.4 Hz, J = 21.2 Hz, CH3), 2.07 (d, 3H, J = 1.6 Hz, Ph-CH3), 2.30
(s, 3H, Ph-CH3), 2.70 (s, 3H, CH3), 4.12-4.22 (m, 1H, CH), 5.46 (d, 1H,
J = 6.4 Hz, NH), 6.19 (s, 1H, Ph-CH), 6.72-6.87 (m, 3H, Ph-H), 7.05-7.11
(m, 4H, Ph-H) (CDCl3)
1.15 (dd, 6H, J = 6.4 Hz, J = 22.8 Hz, CH3), 2.06 (s, 3H, Ph-CH3), 2.27
(s, 3H, Ph-CH3), 2.71 (s, 3H, CH3), 4.13-4.23 (m, 1H, CH), 5.49
(d, 1H, J = 7.2 Hz, NH), 6.18 (s, 1H, Ph-CH), 6.72-6.87 (m, 3H, Ph-H),
6.96-7.14 (m, 4H, Ph-H) (CDCl3)
1.17 (dd, 6H, J = 6.4 Hz, J = 23.6 Hz, CH3), 2.02 (s, 3H, Ph-CH3),
2.49 (s, 3H, Ph-CH3), 2.70 (s, 3H, CH3), 4.16-4.25 (m, 1H, CH),
5.36 (d, 1H, J = 7.2 Hz, NH), 6.57 (s, 1H, Ph-CH), 6.68-7.19
(m, 7H, Ph-H) (CDCl3)
1.15 (dd, 6H, J = 6.4 Hz, J = 23.2 Hz, CH3), 2.07 (d, 3H, J = 1.6 Hz, Ph-CH3),
2.70 (s, 3H, CH3), 4.13-4.22 (m, 1H, CH), 5.50 (d, 1H, J = 6.4 Hz, NH),
5.56 (s, 1H, OH), 6.16 (s, 1H, Ph-CH), 6.71 (d, 2H, J = 8.4 Hz, Ph-H),
6.76-6.86 (m, 3H, Ph-H), 7.07 (d, 2H, J = 8.8 Hz, Ph-H) (CDCl3)
1.17 (dd, 6H, J = 6.0 Hz, J = 25.2 Hz, CH3), 1.98 (s, 3H, Ph-CH3), 2.58
(s, 3H, CH3), 3.95-3.98 (m, 1H, CH), 6.21 (s, 1H, Ph-CH), 6.53-7.01
(m, 7H, Ph-H), 8.17 (br, 1H, NH), 9.34 (s, 1H, OH) (DMSO-d6)
1.22 (dd, 6H, J = 6.4 Hz, J = 13.6 Hz, CH3), 2.09 (d, 3H, J = 1.2 Hz, Ph-CH3),
2.71 (s, 3H, CH3), 4.15-4.24 (m, 1H, CH), 6.19 (s, 1H, Ph-CH), 6.21
(s, 1H, NH), 6.71-7.21 (m, 7H, Ph-H) (CDCl3)
II4
65
white solid, 148-169
[M þ Naþ]: 494.1274;
found: 494.1262
II5
II6
II7
II8
63
55
63
54
white solid, 147-148
white solid, 171-172
white solid, 173
[M þ Naþ]: 483.1034;
found: 483.1034
[M þ Naþ]: 483.1034;
found: 483.1033
[M þ Naþ]: 483.1034;
found: 483.1031
white solid, 145-146
[M þ Naþ]: 467.1329;
found: 467.1329
II9
56
75
57
white solid, 162-163
white solid, 173-174
white solid, 173-174
[M þ Naþ]: 467.1329;
found: 467.1325
II10
[M þ Naþ]: 467.1329;
found: 467.1325
II11
[M þ Naþ]: 517.1297;
found: 517.1286
II12
53
55
white solid, 144-145
white solid, 147-148
[M þ Naþ]: 517.1297;
found: 517.1293
II13
[M þ Naþ]: 517.1297;
found: 517.1289
II14
II15
II16
II17
50
63
47
48
white solid, 171-172
white solid, 166-167
white solid, 201-202
white solid, 160-161
[M þ Naþ]: 463.1580;
found: 463.1581
[M þ Naþ]: 463.1580;
found: 463.1578
[M þ Naþ]: 463.1580;
found: 463.1571
[M þ Naþ]: 465.1373;
found: 465.1375
II18
II19
III1
64
6
white solid, 202-203
white solid, 154-156
white solid, 173-174
[M þ Naþ]: 465.1373;
found: 465.1368
[M þ Naþ]: 465.1373;
found: 465.1366
73
1.02-1.97 (m, 10H, CH3), 2.05 (s, 3H, Ph-CH3), 2.70 (s, 3H, CH3), 3.85-3.87
(m, 1H, CH), 5.56 (d, 1H, J = 7.6 Hz, NH), 6.26 (s, 1H, Ph-CH), 6.71-6.86
(m, 3H, Ph-H), 7.22-7.25 (m, 5H, Ph-H) (CDCl3)
[M þ Naþ]: 489.1736;
found: 487.1732)

7850 J. Agric. Food Chem., Vol. 58, No. 13, 2010
Table 1. Continued
Zheng et al.
compd yield (%) appearance and mp (°C)
1H NMR, δ (solvent)
HRMS (m/z)
III2
90
white solid, 181-182
white solid, 183-184
1.06-2.02 (m, 10H, CH3), 2.07 (s, 3H, Ph-CH3), 2.71 (s, 3H, CH3), 3.85-3.92
(m, 1H, CH), 5.85 (d, 1H, J = 7.6 Hz, NH), 6.27 (s, 1H, Ph-CH), 6.70-6.90
(m, 3H, Ph-H), 7.48 (d, 2H, J = 8.4 Hz, Ph-H), 8.12 (d, 2H, J = 8.4 Hz,
Ph-H) (CDCl3)
1.09-2.03 (m, 10H, CH3), 2.07 (s, 3H, Ph-CH3), 2.72 (s, 3H, CH3), 3.86-3.93
(m, 1H, CH), 5.92 (d, 1H, J = 7.2 Hz, NH), 6.31 (s, 1H, Ph-CH), 6.72-6.85
(m, 3H, Ph-H), 7.45 (t, 1H, J = 8.0 Hz, Ph-H), 7.63 (d, 1H, J = 7.6 Hz, Ph-H),
8.13, 8.15 (d, 2H, Ph-H) (CDCl3)
1.10-1.99 (m, 10H, CH3), 2.05 (s, 3H, Ph-CH3), 2.74 (s, 3H, CH3), 3.81-3.89
(m, 1H, CH), 5.88 (d, 1H, J = 7.2 Hz, NH), 6.78 (s, 1H, Ph-CH), 6.79-6.91
(m, 3H, Ph-H), 7.43-7.95 (m, 4H, Ph-H) (CDCl3)
1.04-1.99 (m, 10H, CH3), 2.08 (s, 3H, Ph-CH3), 2.70 (s, 3H, CH3), 3.82-3.89
(m, 1H, CH), 5.58 (d, 1H, J = 7.6 Hz, NH), 6.68 (s, 1H, Ph-CH), 6.78-6.89
(m, 3H, Ph-H), 7.17 (d, 2H, J = 8.4 Hz, Ph-H), 7.24 (d, 2H, J = 8.4 Hz,
Ph-H) (CDCl3)
1.07-2.00 (m, 10H, CH3), 2.08 (s, 3H, Ph-CH3), 2.71 (s, 3H, CH3), 3.83-3.91
(m, 1H, CH), 5.68 (d, 1H, J = 7.6 Hz, NH), 6.19 (s, 1H, Ph-CH), 6.73-6.88
(m, 3H, Ph-H), 7.10-7.27 (m, 4H, Ph-H) (CDCl3)
1.07-2.04 (m, 10H, CH3), 2.01 (s, 3H, Ph-CH3), 2.68 (s, 3H, CH3), 3.87-3.94
(m, 1H, CH), 5.62 (d, 1H, J = 7.6 Hz, NH), 6.66 (s, 1H, Ph-CH), 6.71-6.95
(m, 3H, Ph-H), 7.02-7.36 (m, 4H, Ph-H) (CDCl3)
1.04-1.99 (m, 10H, CH3), 2.07 (s, 3H, Ph-CH3), 2.70 (s, 3H, CH3), 3.85-3.87
(m, 1H, CH), 5.58 (d, 1H, J = 7.2 Hz, NH), 6.24 (s, 1H, Ph-CH), 6.68-7.22
(m, 7H, Ph-H) (CDCl3)
1.06-1.99 (m, 10H, CH3), 2.07 (s, 3H, Ph-CH3), 2.70 (s, 3H, CH3), 3.83-3.90
(m, 1H, CH), 5.67 (d, 1H, J = 7.2 Hz, NH), 6.19 (s, 1H, Ph-CH), 6.73-6.86
(m, 3H, Ph-H), 6.96-7.25 (m, 4H, Ph-H) (CDCl3)
1.06-2.00 (m, 10H, CH3), 2.04 (s, 3H, Ph-CH3), 2.70 ( s, 3H, CH3), 3.86-3.93
(m, 1H, CH), 5.69 (d, 1H, J = 7.2 Hz, NH), 6.51 (s, 1H, Ph-CH), 6.75-7.24
(m, 7H, Ph-H) (CDCl3)
1.06-2.00 (m, 10H, CH3), 2.08 (s, 3H, Ph-CH3), 2.70 (s, 3H, CH3), 3.84-3.91
(m, 1H, CH), 5.70 (d, 1H, J = 7.2 Hz, NH), 6.23 (s, 1H, Ph-CH), 6.69-6.88
(m, 3H, Ph-H), 7.40 (d, 2H, J = 8.0 Hz, Ph-H), 7.53 (d, 2H, J = 8.0 Hz,
Ph-H) (CDCl3)
1.05-2.00 (m, 10H, CH3), 2.06 (s, 3H, Ph-CH3), 2.71 (s, 3H, CH3), 3.85-3.92
(m, 1H, CH), 5.70 (d, 1H, J = 7.6 Hz, NH), 6.31 (s, 1H, Ph-CH), 6.67-6.81
(m, 3H, Ph-H), 7.37-7.54 (m, 4H, Ph-H) (CDCl3)
0.99-1.85 (m, 10H, CH3), 2.04 (s, 3H, Ph-CH3), 2.71 (s, 3H, CH3), 3.82-3.89
(m, 1H, CH), 5.40 (d, 1H, J = 7.6 Hz, NH), 6.64 (s, 1H, Ph-CH), 6.73-7.24
(m, 5H, Ph-H), 7.36 (t, 1H, J = 7.6 Hz, Ph-H), 7.70 (d, 1H, J = 8.0 Hz,
Ph-H) (CDCl3)
1.02-1.96 (m, 10H, CH3), 2.06 (s, 3H, Ph-CH3), 2.30 (s, 3H, Ph-CH3), 2.70
(s, 3H, CH3), 3.84-3.86 (m, 1H, CH), 5.52 (d, 1H, J = 7.6 Hz, NH), 6.22
(s, 1H, Ph-CH), 6.71-6.86 (m, 3H, Ph-H), 7.05 (d, 2H, J = 8.0 Hz, Ph-H),
7.09 (d, 2H, J = 8.0 Hz, Ph-H) (CDCl3)
1.03-1.97 (m, 10H, CH3), 2.06 (s, 3H, Ph-CH3), 2.27 (s, 3H, Ph-CH3), 2.71
(s, 3H, CH3), 3.83-3.90 (m, 1H, CH), 5.54 (d, 1H, J = 7.6 Hz, NH), 6.22
(s, 1H, Ph-CH), 6.70-6.86 (m, 3H, Ph-H), 6.95-7.14 (m, 4H,
Ph-H) (CDCl3)
1.03-1.97 (m, 10H, CH3), 2.01 (s, 3H, Ph-CH3), 2.49 (s, 3H, Ph-CH3),
2.70 (s, 3H, CH3), 3.85-3.93 (m, 1H, CH), 5.41 (d, 1H, J = 7.2 Hz, NH),
6.60 (s, 1H, Ph-CH), 6.69-7.19 (m, 7H, Ph-H) (CDCl3)
1.03-1.96 (m, 10H, CH3), 2.06 (s, 3H, Ph-CH3), 2.70 (s, 3H, CH3),
3.84-3.86 (m, 1H, CH), 5.58 (d, 1H, J = 8.0 Hz, NH), 5.94 (d, 1H,
J = 18.0 Hz, OH), 6.18 (s, 1H, Ph-CH), 6.71 (d, 2H, J = 8.4 Hz, Ph-H),
6.75-6.85 (m, 3H, Ph-H), 7.06 (d, 2H, J = 8.4 Hz, Ph-H) (CDCl3)
1.05-1.95 (m, 10H, CH3), 2.06 (s, 3H, Ph-CH3), 2.62 (s, 3H, CH3), 3.80-3.88
(m, 1H, CH), 6.07 (s, 1H, OH), 6.18 (s, 1H, Ph-CH), 6.61 (d, 1H, J = 7.6 Hz, NH),
6.75-7.23 (m, 7H, Ph-H) (CDCl3)
[M þ Naþ]: 534.1587;
found: 534.1575
III3
98
[M þ Naþ]: 534.1587;
found: 534.1577
III4
III5
72
97
yellow solid, 113-115
white solid, 180-181
[M þ Naþ]: 534.1587;
found: 534.1579
[M þ Naþ]: 523.1347;
found: 523.1339
III6
77
82
92
72
60
85
white solid, 178-179
white solid, 176
[M þ Naþ]: 523.1347;
found: 523.1333
III7
[M þ Naþ]: 523.1347;
found: 523.1333
III8
white solid, 178
[M þ Naþ]: 507.1642;
found: 507.1635
III9
white solid, 176-177
white solid, 169-170
white solid, 176-177
[M þ Naþ]: 507.1642;
found: 507.1637
III10
III11
[M þ Naþ]: 507.1642;
found: 507.1633
[M þ Naþ]: 557.1610;
found: 557.1611
III12
III13
39
84
white solid, 160-161
white solid, 83-85
[M þ Naþ]: 557.1610;
found: 557.1610
[M þ Naþ]: 557.1610;
found: 557.1597
III14
90
65
white solid, 170
white solid, 170
[M þ Naþ]: 503.1893;
found: 503.1892
III15
[M þ Naþ]: 503.1893;
found: 503.1889
III16
III17
51
50
white solid, 178-179
white solid, 148-149
[M þ Naþ]: 503.1893;
found: 508.1882
[M þ Naþ]: 505.1686;
found: 505.1672
III18
27
white solid, 183-184
[M þ Naþ]: 505.1686;
found: 505.1680
This study used U-4CR as the synthesis method via parallel
synthesis, and the target compounds were proved to be easily
synthesized and purified. Following the method developed in our
previous study (12), we mixed an amine component and an
aldehyde component first to get an intermediate imine, and then
a carbonyl component was added in the mixture for certain time
intervals before the addition of an isocyanide component. The
amount of methanol added to the reaction was as small as
possible, just enough to make the agitation effective. After all
components were mixed, products usually precipitated in solvent
within 1 h. To obtain high yields, we removed solvent under
reduced pressure and then purified the products by column
chromatography on silica gel using ethyl acetate and petroleum
ether (60-90 °C) at 1:3 as an eluent. Generally, compounds in

Article
J. Agric. Food Chem., Vol. 58, No. 13, 2010 7851
Table 2. Fungal Growth Inhibition by the Target Compounds (Percent)a
Scheme 1. Molecule Structures and Synthesis of the Target Compoundsa
compd AS BC CA CB CL FO GZ PI PP PS PT RS
I1
39 85 21
39 79 17
0
9
11 36 13 12 47 64 nd nd
23 21 33 60 nd nd
I2
4
4
I3
33 78
26 60
9
5
46 10 12 33 26 45 52 nd nd
a Series I: X = 3-chloro-4-methylphenyl, Z = isopropyl, Y = phenyl (I1);
4-nitrophenyl (I2); 3-nitrophenyl (I3); 4-nitrophenyl (I4); 4-chlorophenyl (I5); 3-chloro-
phenyl (I6); 2-chlorophenyl (I7); 4-fluorophenyl (I8); 3-fluorophenyl (I9); 4-fluoro-
phenyl (I10); 4-(trifluoromethyl)phenyl (I11); 3-(trifluoromethyl)phenyl (I12); 2-(trif-
luoromethyl)phenyl (I13); 4-methylphenyl (I14); 3-methylphenyl (I15); 2-methylphenyl
(I16); 4-hydroxyphenyl (I17); 3-hydroxyphenyl (I18); 2-hydroxyphenyl (I19). Series II:
X = 3-fluoro-4-methylphenyl, Z = isopropyl, Y = phenyl (II1); 4-nitrophenyl (II2);
3-nitrophenyl (II3); 4-nitrophenyl (II4); 4-chlorophenyl (II5); 3-chlorophenyl (II6);
2-chlorophenyl (II7); 4-fluorophenyl (II8); 3-fluorophenyl (II9); 4-fluorophenyl (II10);
4-(trifluoromethyl)phenyl (II11); 3-(trifluoromethyl)phenyl (II12); 2-(trifluoromethyl)-
phenyl (II13); 4-methylphenyl (II14); 3-methylphenyl (II15); 2-methylphenyl (II16);
4-hydroxyphenyl (II17); 3-hydroxyphenyl (II18); 2-hydroxyphenyl (II19). Series III: X =
3-fluoro-4-methylphenyl, Z = cyclohexyl, X = 3-fluoro-4-methylphenyl, Z = isopropyl,
Y = phenyl (III1); 4-nitrophenyl (III2); 3-nitrophenyl (III3); 4-nitrophenyl (III4);
4-chlorophenyl (III5); 3-chlorophenyl (III6); 2-chlorophenyl (III7); 4-fluorophenyl
(III8); 3-fluorophenyl (III9); 4-fluorophenyl (III10); 4-(trifluoromethyl)phenyl (III11);
3-(trifluoromethyl)phenyl (III12); 2-(trifluoromethyl)phenyl (III13); 4-methylphenyl
(III14); 3-methylphenyl (III15); 2-methylphenyl (III16); 4-hydroxyphenyl (III17);
3-hydroxyphenyl (III18).
I4
8
9
9
10 14 12 24 39 31 nd nd
I5
37 76 21
37 76
7
14 26
26
4
4
19 47 60 nd nd
14 40 62 nd nd
I6
8
I7
49 76 21 25 13 23 23 23 27 72 nd nd
42 70 13 25 17 14 27 36 63 nd nd
I8
5
I9
42 78 21 33 10 19 23 18 38 69 nd nd
40 73 21 25 10 21 37 15 33 67 nd nd
I10
I11
I12
I13
I14
I15
I16
I17
I18
I19
II1
35 70 25 17
37 76 13 25 17 14 50
7
12 67
0
0
11 56 nd nd
44 26 nd nd
36 70 18 23 17 10 24 13 43 57 nd nd
34 79 33
44 76 38 17 13 14 37
9
11 19
0
0
8
37 60 nd nd
51 63 nd nd
40 76 21 17 10 19 37 10 31 69 nd nd
37 76 17 17 10 23 20 23 36 65 nd nd
33 73 14 23 13 30 15 30 45 59 nd nd
38 75
13 nd
9
9
0
0
7
18 18 24 39 47 nd nd
14 nd 27 38
nd 13
13 nd 27 25 29 nd 45 13
0
0
0
0
22
0
0
0
2
II2
0
nd 27 25
0
0
17
II3
14 15
hydroxyl at the 2-position interfered with the interaction between
amine and aldehyde.
II4
0 nd
38 nd
0
0
0
0
0
25 14 nd 27
0
38
0
19 39
0
0
0
0
0
0
0
0
2
15
22
0
II5
13 14 nd
0
0
0
0
0
0
0
0
0
0
8
0
0
0
0
0
0
28
4
Biological Activity and Structure-Activity Relationship Study.
Fungicidal Activity. The 12 fungi representing typical fungi often
occurring in the Chinese agro-ecosystem including AS, BC, CA,
CB, CL, FO, GZ, PI, PP, PS, PT, and RS were chosen for
fungicide screening, and the results are shown in Table 2. All
compounds showed a certain degree of fungicidal activity at
50 μg/mL. The results indicated that compounds belonging to
series I showed high activities against BC and PS; most of them
had inhibition activities above 70% against BC and 60% against
PS, respectively. Especially, compound I1 had an inhibition acti-
vity of 85% against BC, and compound III8 had inhibition acti-
vity of 87% against PS. Moreover, compound III1 had inhibition
activities of 69 and 65% against PP and PS, respectively. As the
results describe above, most of the target compounds showed a
very narrow spectrum against fungi tested; however, compound
III10 showed a broad spectrum of activities against several fungi
tested with inhibition activities of 55, 63, 73, 57, 59, and 50%
against CA, AS, GZ, CL, RS, and PI, respectively. These results
indicated that F- or CF3-containing derivatives had good activity
because of their special atom size and their characteristics with
H to form a H bond with the potential target.
Structure and Fungicide Activity Relationship Study. All 56
5-methyl-1,2,3-thiadiazole derivatives with an R-amino acyl-
amide skeleton synthesized were tested by fungicidal activity
and direct antivirus activity in vitro screening. All of the results
provided useful information to study the structure-activity
relationship of these new structures shown in Scheme 1. Herein,
the effect of substitutions at positions X and Y (Scheme 1) on the
bioactivity were studied, respectively. At position X, we paid
attention to the comparison of 3-chloro-4-methylphenyl with
3-fluoro-4-methylphenyl. At position Y, the substitutions at the
substituted phenyl were compared with each other. Finally, at
position Z, we compared cyclohexyl with isopropyl. Only significant
effects are mentioned; if not, there was no significant structure-
-activity relationship observed on the bioactivity concerned.
For fungi PP, as described in Figure 1, at position X, com-
pounds with 3-chloro-4-methylphenyl showed significantly better
activity thanthe others. This indicated that a Cl atomhere was the
key factor favoring the bioactivity. For fungi PS, as described
in Figure 2, at position X, compounds with 3-chloro-4-methyl-
phenyl showed significantly better activity than the others.
II6
0
0
0
nd
nd
nd
13
0
0 nd
14 nd
II7
25
0
0
II8
25
0
nd 27
0
22
43
15
15
II9
44 nd 27 25 57 nd
5
13 14 nd 27 44
19
0
II10
II11
II12
II13
II14
II15
II16
II17
II18
II19
III1
III2
III3
III4
III5
III6
III7
III8
III9
III10
III11
III12
III13
III14
III15
III16
III17
III18
TDL
38 nd
0
0
0
0
0
nd
25
0
nd
0
25
11
4
nd 27 25 14 nd 36
0
13 36 nd 27 44
14 22
17
11 14 37
13 nd
31 nd 45 25 29 nd
9
15
0
0
0
13
13
0
nd
9
56 nd 36
25
0
0
nd
0
0
4
0
0
0
0
0
0
0
0
0
0
0
0
22
0
50 nd 27 31
0
0
0
nd
nd
nd
0
0
0
0
0
nd
11 14 nd
0
0
0
38
0
2
0
0
50 nd 50
0
19 41
26
0
25 nd 18
nd 27 38 69 65
0
0
0
0
nd 45 25 14 nd
0
0
0
0
50
44
0
0
0
0
2
nd
0
25
0
0
0
0
0
nd
nd
nd
4
nd 27 25
nd 18 13
50 12
0
9
6
0
8
0
0
8
0
0
0
0
8
0
0
0
0
0
22
25 nd 27 25
nd 27 44
nd 45 25
0
14 39
0
0
nd
nd
0
9
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
22
2
14 nd
0
9
25
44
87
11
0
50 nd 27 25 43 nd
2
63 nd 55 25 57 nd 73 50
nd 14 nd
25 nd 27 25 29 nd 14 25
59
22
22
4
0
0
0
9
0
0
13
0
44 nd
0
nd 27 25 14 nd
25 14 nd
0
0
25
25
1
0
6
15
0
2
nd
0
0
25
0
25 29 nd
nd 36
0
37
22
22
30
1
38 nd
44
0
13 nd 45 25 36 nd 14 25
nd 25 nd 14
0
0
0
0
6
0
nd 67 18 40 nd 48 36 46 74 46 nd
a nd, not detected; AS, Alternaria solani; BC, Botrytis cinerea; CA, Cercospora
arachidicola; CB, Cercospora beticola; CL, Colletotrichum lagenarium; FO, Fusar-
ium oxysporum; GZ, Gibberella zeae; PI, Phytophthora infestans (Mont) de Bary;
PP, Physalospora piricola; PS, Pellicularia sasakii; PT, Puccinia triticina Eriks; RS,
Rhizoctonia solani; TDL, tiadinil.
series III had higher yields; this indicated that using cyclohexyl
isocyanide as the isocyanide component could favor the reaction.
Moreover, when 2-hydroxybenzaldehyde served as the carbonyl
compound, the yields were pretty low, which indicated that the

7852 J. Agric. Food Chem., Vol. 58, No. 13, 2010
Zheng et al.
Figure 1. Parallel activity contrasts between series I, series II, and series III against PP. The inhibition activity was tested at 50 μg/mL and expressed in
ordinate. The figures under the X-axis represent the substitutions at position Y in Scheme 1: 1 = phenyl; 2 = 4-nitrophenyl; 3 = 3-nitrophenyl; 4 = 4-nitrophenyl;
5 = 4-chlorophenyl; 6 = 3-chlorophenyl; 7 = 2-chlorophenyl; 8 = 4-fluorophenyl; 9 = 3-fluorophenyl; 10 = 4-fluorophenyl; 11 = 4-(trifluoromethyl)phenyl; 12 =
3-(trifluoromethyl)phenyl; 13 = 2-(trifluoromethyl)phenyl; 14 = 4-methylphenyl; 15 = 3-methylphenyl; 16 = 2-methylphenyl; 17 = 4-hydroxyphenyl; 18 =
3-hydroxyphenyl; 19 = 2-hydroxyphenyl. Compounds at the same abscissa had the same substitutions at position Y inScheme 1 and compounds belonged to
the same series had the same substitutions at position X in Scheme 1.
Figure 2. Parallel activity contrasts between series I, series II, and series III againstPS. The inhibition activities were tested at 50μg/mL. The figures underthe
abscissa are as explained in Figure 1.
Figure 3. Parallel activity contrasts between series I, series II, and series III againstCL. The inhibition activities were tested at 50 μg/mL. The figures under the
abscissa are as explained in Figure 1.

Article
J. Agric. Food Chem., Vol. 58, No. 13, 2010 7853
Figure 4. Parallel activity contrasts betweenseries I, series II, andseries III againstRS. The inhibition activities were testedat50μg/mL. The figures underthe
abscissa are the same as explained in Figure 1.
Figure 5. Parallel activity contrasts between series I, series II, and series III against PI. The inhibition activities were tested at 50 μg/mL. The figures under the
abscissa are the same as explained in Figure 1.
This indicated that a Cl atom here was the key factor making
contribution to the bioactivity. At position Y, compounds with
phenyl and 4-fluorophenyl were better in most contrasts. For
fungi CL, as described in Figure 3, at position X, compounds with
3-fluoro-4-methylphenyl showed significantly better activity than
the others. This indicated that a F atom here was the key factor
favoring the bioactivity. For fungi RS, as described in Figure 4, at
position Z, compounds with isopropyl showed significantly better
activity than the others. For fungi PI, as described in Figure 5, at
position Z, compounds with cyclohexyl showed significantly
better activity than the others.
Anti-TMV Activity in Vitro and in Vivo. TDL and ribavirin
were used as two positive controls. The results of activity screen-
ing by half-leaf juice-robbing method against TMV indicated
that (Table 3) most compounds showed a certain degree of
direct inhibition activities against TMV in vitro with results
lower than 50%. However, some compounds exhibited relatively
higher activity; compounds II6, III2, and III12 had direct inhibition
activities of 66, 70, and 72%, respectively, at 500 μg/mL, which were
over twice as much as ribavirin did. They did not keep the high
activity at 100 μg/mL. Only III1 and II13 had high activity at both
500 and 100 μg/mL. Their activities were >70% at 500 μg/mL and
>50% at 100 μg/mL, significantly higher than those of the positive
control agents. Compounds III1 and II13 are worthy of further
study.
Moreover, 16 compounds were chosen for further evaluation of
protection, inactivation, and curative and induction activities
against TMV in vivo. The results in Table 4 indicate that almost
all of the compounds tested had good inactivation activity. Among
these, compounds I15, II10, III6, andIII8 had inactivation activities of
82, 74, 79, and 79%, respectively, at 500 μg/mL, and 72, 73, 69, and
69%, respectively, at 100 μg/mL, which were significantly better
than the positive control agents. As for the protection, curative
effect, and induction activities, most of them had lower activities
than the positive control. However, some compounds stood out.
Compound II17 had good protection activities of 76 and 71% at
500 and 100 μg/mL, respectively, and compound I10 had curative
activities of 60 and 47% at 500 and 100 μg/mL, respectively, which
were equal to or higher than that of the positive control. These
results indicated that the title compounds had good potential of
antifungal and anti-TMV bioactivities. There was observed no
phytotoxicity to tobacco during the test courses of experiment.
Structure and Anti-TMV Activity Relationship Studies. No
significant structure-activity relationship was observed. This
indicated that the whole molecular structure played an important
part in these activities rather than one moiety in the molecule.

7854 J. Agric. Food Chem., Vol. 58, No. 13, 2010
Table 3. Inhibition Activities against TMV in Vitro
Zheng et al.
compd
concn (μg/mL)
activity (%)
compd
concn (μg/mL)
activity (%)
compd
III1
concn (μg/mL)
activity (%)
I1
200
500
200
500
100
500
100
500
200
500
200
500
200
500
200
500
100
500
100
500
200
500
200
500
200
500
200
500
200
500
200
500
200
500
100
500
100
500
100
500
30 ( 1
43 ( 3
14 ( 1
19 ( 3
28 ( 11
34 ( 13
19 ( 4
59 ( 13
25
II1
100
500
100
500
100
500
100
500
100
500
100
500
100
500
100
500
100
500
100
500
100
500
100
500
100
500
100
500
100
500
100
500
100
500
100
500
100
500
100
500
5 ( 4
15 ( 5
9 ( 8
25 ( 11
4 ( 4
14 ( 5
11 ( 10
24 ( 10
20 ( 8
12 ( 2
32 ( 2
66 ( 5
23 ( 5
17 ( 2
18 ( 9
51 ( 1
50
100
500
100
500
100
500
100
500
100
500
100
500
100
500
100
500
100
500
100
500
100
500
100
500
100
500
100
500
100
500
100
500
100
500
100
500
51
71 ( 3
8 ( 10
70 ( 4
9 ( 10
57 ( 4
5 ( 8
I2
II2
III2
I3
II3
III3
I4
II4
III4
11 ( 2
8 ( 1
I5
II5
III5
32 ( 5
6 ( 1
9 ( 1
3 ( 5
8 ( 1
14 ( 3
19 ( 7
34 ( 4
18 ( 3
32 ( 5
34 ( 5
32 ( 5
34 ( 5
17
14 ( 2
14 ( 6
18 ( 9
16 ( 3
53 ( 7
24 ( 2
44 ( 5
52 ( 1
30 ( 2
33 ( 1
58 ( 1
27 ( 6
36 ( 4
35 ( 1
72 ( 11
31 ( 6
20 ( 5
21 ( 1
53 ( 11
44 ( 6
29 ( 2
13 ( 3
35 ( 5
9 ( 2
I6
II6
III6
I7
II7
III7
I8
II8
III8
I9
II9
III9
30
I10
I11
I12
I13
I14
I15
I16
I17
I18
I19
ribavirin
II10
II11
II12
II13
II14
II15
II16
II17
II18
II19
tiadinil
16 ( 9
46 ( 2
14 ( 2
45 ( 2
19 ( 1
51 ( 3
57 ( 1
71 ( 5
7 ( 3
31 ( 3
14 ( 3
35 ( 1
36 ( 8
55 ( 1
26 ( 4
35 ( 1
4 ( 4
III10
III11
III12
III13
III14
III15
III16
III17
III18
41
29
16
11
21
18 ( 3
34 ( 4
58
12
0
4
34 ( 9
25 ( 2
48 ( 1
13 ( 8
24 ( 2
18 ( 4
23 ( 2
20 ( 2
32 ( 1
20 ( 5
27 ( 7
56 ( 1
25 ( 2
48 ( 1
Table 4. Antivirus in Vivo and Systemic Acquired Resistance Activity of the Title Compounds (Percent)
protective effect (%) curative effect (%) inactivation effect (%)
induction effect (%)
compd
500 μg/mL
100 μg/mL
500 μg/mL
100 μg/mL
500 μg/mL
100 μg/mL
100 μg/mL
50 μg/mL
I1
57 ( 5
48 ( 5
38 ( 10
36 ( 2
27 ( 10
60 ( 6
44 ( 5
76 ( 5
67
40 ( 2
14 ( 5
13 ( 6
8 ( 10
26 ( 7
27 ( 7
5 ( 4
71 ( 0
29 ( 8
6 ( 7
46 ( 7
40 ( 8
42 ( 7
nd
30 ( 4
33 ( 4
21 ( 3
nd
65 ( 12
82 ( 4
50 ( 4
64 ( 5
62 ( 8
74 ( 9
65 ( 4
72 ( 4
79 ( 4
79 ( 4
69 ( 8
67 ( 4
59 ( 4
59 ( 9
50 ( 4
72 ( 4
23 ( 8
50 ( 4
65 ( 4
73 ( 4
67 ( 9
51 ( 9
69
41 ( 6
49 ( 8
0
33 ( 9
65 ( 3
37 ( 7
23 ( 5
66 ( 4
59 ( 3
30 ( 4
36 ( 11
30 ( 5
41 ( 16
40 ( 8
32 ( 9
50 ( 6
31 ( 9
I15
I16
I17
41 ( 3
7 ( 7
7 ( 8
60 ( 6
7 ( 8
22 ( 0
41 ( 7
54 ( 7
76 ( 5
52 ( 4
28 ( 9
II2
II10
17 ( 4
60 ( 6
44 ( 5
55 ( 4
40 ( 8
52 ( 5
38 ( 5
57 ( 2
58 ( 7
nd
0 ( 7
47 ( 4
32 ( 6
28 ( 6
0
II13
II17
III6
III8
17 ( 8
8 ( 3
44 ( 7
75
20 ( 10
29 ( 7
23 ( 2
46 ( 8
nd
69 ( 8
38 ( 8
44 ( 4
42 ( 4
46 ( 8
III15
III18
tiadinil
ribavirin
10 ( 5
43 ( 5
57
59 ( 7
30 ( 7
ABBREVIATIONS USED
magnetic resonance; HRMS, high-resolution mass spectrometry;
nd, not detected; PDA, potato dextrose agar; PG, Phyricularia
grisea; PI, Phytophthora infestans (Mont) de Bary; PP, Physalos-
pora piricola; PS, Pellicularia sasakii; RS, Rhizoctonia solani; SOP,
standard operation practice; TDL, tiadinil; TMS, tetramethylsilane;
AS, Alternaria solani; BTH, acibenzolar-S-methyl; CA, Cercos-
pora arachidicola; CB, Cercospora beticola; CL, Colletotrichum
lagenarium; DMSO-d6, deutero-, dimethyl sulfoxide; FO, Fusarium
oxysporum; GZ, Gibberella zeae; 1H NMR, hydrogen nuclear

Article
J. Agric. Food Chem., Vol. 58, No. 13, 2010 7855
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Received for review February 19, 2010. Revised manuscript received
June 5, 2010. Accepted June 6, 2010. This study was funded in part by
grants from the National Natural Science Foundation of China (No.
20872071), the International Collaboration Program of the National
Natural Science Foundation of China (No. 20911120069), the Tianjin
Natural Science Foundation (10JCZDJC17500), the National Key
Project for Basic Research (2010CB126105), and the Key Laboratory
of Pesticide Chemistry and Application, Ministry of Agriculture
(MOA), Beijing, People’s Republic of China (No. MOAPCA200903).