Discovery of 5-(5,5-Dimethylbutenolide-3-ethylidene)-2-aminoimidazolinone Derivatives as Fungicidal Agents

Article abstract of DOI:10.3390/molecules200813740

The novel fungicidal agents 5-(5,5-dimethylbutenolide-3-ethylidene)-2-aminoimidazolinone derivatives, were designed and synthesized in moderate to excellent yields in four steps by α-hydroxyketone and diketene as raw materials and characterized by HR-ESI-MS and 1H-NMR. The preliminary bioassay showed that some of these compounds, such as 4a, 4e and 5g exhibit 94.9%, 92.8% and 81.4% inhibition rates against Sclerotinia scleotiorum at the concentration of 50 μg/mL, respectively. The EC50 values of compounds 4e and 4i were 4.14 and 3.27 μM against Alternaria Solani, and 5g had EC50 value of 3.23 μM against S. scleotiorum. Compounds 4d and 4g displayed 98.0% and 97.8% control of spore germination against Botrytis cinerea at the concentration of 100 μg/mL, respectively.

Full text of DOI:10.3390/molecules200813740

Molecules 2015, 20, 13740-13752; doi:10.3390/molecules200813740
OPEN ACCESS
molecules
ISSN 1420-3049
www.mdpi.com/journal/molecules
Article
Discovery of 5-(5,5-Dimethylbutenolide-3-ethylidene)-2-amino-
imidazolinone Derivatives as Fungicidal Agents
Bo Tang, Mingyan Yang, Yu Zhao, Lingqing Kong, Weiwei Wang and Mingan Wang *
Department of Applied Chemistry, China Agricultural University, Beijing 100193, China;
E-Mails: bs20143100240@cau.edu.cn (B.T.); yangmy@cau.edu.cn (M.Y.); lizhl@cau.edu.cn (Y.Z.);
klq36211314@cau.edu.cn (L.K.); jay2hebe@cau.edu.cn (W.W.)
* Author to whom correspondence should be addressed; E-Mail: wangma@cau.edu.cn;
Tel.: +86-10-6273-4093.
Academic Editor: Jean Jacques Vanden Eynde
Received: 3 July 2015 / Accepted: 24 July 2015 / Published: 28 July 2015
Abstract: The novel fungicidal agents 5-(5,5-dimethylbutenolide-3-ethylidene)-2-amino-
imidazolinone derivatives, were designed and synthesized in moderate to excellent yields in
four steps by α-hydroxyketone and diketene as raw materials and characterized by HR-ESI-MS
and 1H-NMR. The preliminary bioassay showed that some of these compounds, such as 4a,
4e and 5g exhibit 94.9%, 92.8% and 81.4% inhibition rates against Sclerotinia scleotiorum
at the concentration of 50 µg/mL, respectively. The EC50 values of compounds 4e and 4i
were 4.14 and 3.27 µM against Alternaria Solani, and 5g had EC50 value of 3.23 µM
against S. scleotiorum. Compounds 4d and 4g displayed 98.0% and 97.8% control of spore
germination against Botrytis cinerea at the concentration of 100 µg/mL, respectively.
Keywords: butenolide; 2-aminoimidazolinone; synthesis; fungicidal activity
1. Introduction
In recent years, various imidazolinone-containing compounds have been found in nature or
synthesized and reported to display a wide range of biological activity such as fenamidone (Scheme 1)
having antimicrobial and fungicidal activities [1–9]. Several 5-cyclohexylidene-2-aminoimidazolin-4-
one derivatives have been prepared in our laboratory and showed significant fungicidal activities against
several important phytopathgens [10,11]. Furthermore, the natural and unnatural butenolide derivatives

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including 3-acetyl-5,5-dimethylbutenolide (Scheme 1) exhibited interesting biological activities [12–16].
Some heterocyclic compounds derived from 3-acetyl-5,5-dimethylbutenolide were also reported,
but their biological activities were not disclosed [17,18]. In order to continue our exploration of novel
biologically active 2-aminoimidazolin-4-one heterocyclic compounds [10,11,19], butenolide and
2-aminoimidazolin-4-one were combined into a new class of 2-aminoimidazolin-4-one derivatives
containing 5,5-dimethylbutenolide heterocycle derivatives as inhibitors of mitochondrial respiration,
(Schemes 1 and 2) to improve their activities against the important agricultural diseases, which had
not been published in literatures. In this paper, a series of 5-(5,5-dimethylbutenolide-3-ethylidene)-2-
aminoimidazolinone derivatives were designed and synthesized, and their fungicidal activities and the
structure-activity relationships were explored.
Scheme 1. The design of target molecules based on the active groups.
Scheme 2. The synthetic route of 5-(butenolide-3-ethylidene)-2-aminoimidazolinone derivatives.
2. Results and Discussion
As indicated in the introduction, both imidazolinone and butenolide derivatives displayed a wide
range of biological activity [1–9,12–16]. Based on these characters, we combined the butenolide and
2-aminoimidazolin-4-one into one molecule (Scheme 1) and synthesized the novel 2-aminoimidazolin-4-one
derivatives containing butenolide heterocycles (4, 5). As shown in Scheme 2, the intermediates 1a and 1b
were synthesized by the reaction of α-hydroxyketone with diketene in 80% and 84% yields following the
literature procedure [20], followed by reacting with thiohydantion catalysed by 2-aminoethanol to afford 2a
and 2b in 63% and 67% yields, respectively [10,11]. Then methylations of 2a and 2b with methyl iodide
were carried out at ambienent temperature to give the key intermediates 3a and 3b in 90% and 79%
yields [19]. Finally, the intermediates 3a and 3b reacted with anilines or benzyl amines under reflux to

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produce the target compounds 4a–4t and 5a–5l in moderate to excellent yields. Interestingly, if acetic
acid was used as solvent, compound 6 was isolated as the byproduct in 10%–40% yield when we first
run the preparing reactions of 4a–4t and fully characterized it by 1H-NMR, 13C-NMR and HR-ESI-MS.
However, after changing from acetic acid to dioxane and adding oxalic acid to the reaction system,
the byproduct was not observed. Moreover, purification of the final products was easier and the yields
were also improved. So the preparation of compounds 5a–5l selected the dioxane as solvent and added
1
oxalic acid into the reaction system to effectively avoid the byproduct. In the H-NMR spectrum, the
two methyl groups had the same chemical shifts in compounds 1a, 2a, 3a and 4a–4t, while the two
methyl groups in compounds 1b, 2b, 3b and 5a–5l had different chemical shifts due to the orientation
1
of phenyl group at 4-position in the butenolide moiety (see Supplementary Materials). The H-NMR
spectra of 4a–4t and 5a–5l were similar to those compounds in the literatures [10,19].
Based on the data in Table 1, compounds 4 and 5 showed a broad-spectrum of fungicidal activities
against these six tested phytopathgens. They were found to be particularly active against Sclerotinia
scleotiorum and Rhizoctonia solani, for example, 4a, 4e and 5g exhibited 94.9%, 92.8% (much higher
than the commecialized carbendazim and nearly equal to chlorothalonil and fenamidone) and 81.4%
inhibition rates against S. scleotiorum, 4d, 4g, 4i, 4j, 4m and 4t showed 72.9%, 72.9%, 72.1%, 75.7%,
72.0%, and 72.5% inhibition rates against R. solani at the concentration of 50 µg/mL, respectively.
Table 1. The fungicidal activities (inhibition rate, %) of compounds 4 and 5 at 50 µg/mL.
Compd.
1a
1b
2a
2b
3a
3b
4a
4b
4c
n
R2 R3
S. scleotiorum R. solani
A. solani
0.0
B. cinerea
0.0
P. capsici
0.0
F. graminearum
0.0
0.0
0.0
0.0
0.0
0.0
0.0
0.0
0.0
49.7
3.3
66.8
7.7
42.3
1.5
52.2
36.2
20.0
16.8
60.5
27.2
31.8
75.3
32.2
16.5
69.3
36.2
40.2
21.4
45.5
29.1
48.4
28.6
40.3
39.2
33.3
31.6
69.8
59.3
79.4
34.2
24.0
29.3
16.7
15.5
0.0
31.8
3.9
25.4
0.0
72.7
18.6
68.4
63.5
59.6
72.9
61.2
57.6
72.9
56.4
76.1
75.7
44.6
65.3
72.0
66.5
68.2
48.5
64.9
59.5
51.9
0.0
34.0
10.6
21.1
31.6
38.2
38.6
48.7
0.0
0
0
0
0
0
0
0
0
0
0
0
0
1
1
1
1
1
H
H
94.9
33.1
19.1
32.3
92.8
31.7
65.0
16.5
38.9
22.2
18.7
30.8
52.2
36.9
16.3
21.5
21.7
15.4
50.9
60.3
63.2
60.8
64.8
53.9
55.7
37.0
67.7
22.0
44.6
45.9
53.9
39.0
48.1
45.4
61.2
41.2
H
H
4-CH3
4-F
4d
4e
H
4-OCH3
4-CF3
H
H
4f
CH3
H
36.1
47.7
15.7
0.0
4g
4h
4i
3-CF3
2-CH3
2-F
38.0
0.0
H
H
23.9
31.5
37.4
39.2
22.9
51.3
30.7
38.5
45.1
23.5
4j
H
2-OCH3
2-Cl
33.9
41.8
2.1
4k
4l
H
H 2,6-(CH3)2
4m
4n
4o
4p
4q
4r
H
H
H
H
H
H
H
23.0
0.0
4-OCH3
4-Cl
4-F
5.3
33.6
56.6
27.6
3-CF3
2-F
1

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Table 1. Cont.
Compd.
n
R2
R3
S. scleotiorum
21.1
R. solani
60.4
72.5
7.6
A. solani
B. cinerea P. capsici
F. graminearum
4s
1
H
2-Cl
38.5
42.1
0.0
27.0
41.5
21.3
25.1
25.2
15.2
38.6
48.6
26.3
30.3
6.8
21.3
0.0
13.0
10.1
26.4
22.0
37.1
16.7
24.1
32.6
59.9
40.2
20.0
28.3
15.4
27.7
31.6
95.8
100
4t
1
0
0
0
0
0
0
0
0
0
1
1
1
H 2-OCH3
22.2
5a
H
H
H
H
67.8
27.7
13.3
48.0
20.0
50.4
44.4
45.1
37.1
8.7
5b
4-CH3
4-F
29.4
10.9
0.0
11.6
34.9
0.0
5c
44.6
5d
H 4-OCH3
21.4
6.9
5e
H
H
H
H
4-CF3
3-CF3
2-F
58.1
11.4
11.4
12.9
2.3
31.4
65.7
14.4
37.3
0.0
5f
36.8
5g
81.4
5h
2-Cl
57.8
5i
H 2-OCH3
31.2
1.5
5j
H
H
H
18.3
9.6
5.2
0.0
2.4
5k
4-Cl
29.2
0.0
24.4
3.0
12.6
8.5
13.3
0.0
5l
H 2-OCH3
30.0
6.9
6
41.7
46.1
99.9
100
96.4
70.7
100
100
93.5
40.9
100
4.2
4.2
Chlorothalonil
Carbendazim
Fenamidone
100
99.1
34.7
79.8
81.0
97.8
57.1
71.9
Comparing 2a and 3a with 1a, it seems that the thiohydantoin ring was responsible for higher activity.
Change from thione to methylthio made little contribution to the bioactivity against R. solani and
Phytophthora capsici. Consequently, methylthio was replaced with phenylamino to obtain compound
4a, which exhibited lower activity than 3a against R. solani and much lower activity against P. capsici.
Surprisingly, compound 4a was found to have excellent activity (94.9%) on S. scleotiorum, which is
much higher than the commecialized carbendazim and nearly equal to chlorothalonil and fenamidone.
These results indicated that the introduction of phenylamino at the 2-position of the imidazolinone
changed the activity spectrum. Inspired by this outstanding result, attention was turned to prepartion
and evaluation of more analogous of 4a by replacing substituents attached to the phenyl ring. The
results indicated that with a strong electron-withdrawing group, such as trifluoromethyl (CF3) at the
4-positon (4e), almost equal activity against S. scleotiorum was noted. The introduction of CH3, F and
OCH3 to any position resulted in the remarkable decrease in fungicidal activity. The compounds with
substituents of 2-OCH3, 4-OCH3, 3-CF3 and 2-F on phenyl ring, however, exhibited better inhibition
against R. solani, but still lower than three known products, which seemed that there was not much
correlation between electronic effect and activity. Further optimization by replacing phenyl with benzyl
led to much more decreased activities against all treated target phytopathgens.
As for R1 groups at 4-position of butenolide, the results indicated that the thiohydantoin 2a and
2-methylthioimidazolinone 3a exhibited inhibition against all tested phytopathgens, especially against
R. solani and P. capsici. In contrast, 2b and 3b showed a significant decrease in activity against all six
phytopathgens, indicating that R1 groups at 4-position of butenolide play a crucial role for regulating the
bioactivity. This is probably due to steric hindrance of the phenyl group compared with methyl group.
This conclusion was confirmed by comparison of the inhibition rates of 4a–4t and corresponding 5a–5l.

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The results above indicated that the aniline moiety should be optimized around compounds 4a and
4e and R1 prefers substituent with small size in the structure modification in the future. Based on the
above results, the EC50 values were determined for these compounds with more than 65% inhibition
rates. The equations for linear relationships of the logarithmic values of inhibition rates and the logarithmic
values of concentrations were obtained with good coefficent γ2 as indicated in Table 2. The data in Table 2
indicated that 4e, 4i and 5g have EC50 values less than 5.0 µM against S. scleotiorum and A. Solani, close
or weaker than carbendazim and fenamidone, and the others have EC50 values more than 5.0 µM against
all four phytopathgens. Among these compounds, most compounds did not exhibit the inhibition of spore
germination against S. scleotiorum, R. Solani and A. solani, while compounds 4d and 4g demonstrated a
siginificant control of spore germination against B. cinerea with 98.0% and 97.8% inhibition rates at
the concentration of 100 µg/mL, respectively. The inhibition of spore germination against B. cinerea for
4g was still observed even at a concentration of 10 µg/mL, as shown in Figure 1. The mode of action will
be a topic of the future research. These results indicated that there is the possibility to improvement of
fungicidal activities by modification of chemical structures.
Table 2. The EC50 values of compounds 4 and 5 against different phytopathgens.
Compd.
Fungi
S. scleotiorum
S. scleotiorum
S. scleotiorum
S. scleotiorum
S. scleotiorum
R. solani
Equation
Coefficent γ2
0.966
0.990
0.950
0.978
0.981
0.949
0.972
0.956
0.953
0.971
0.963
0.972
0.981
0.953
0.962
0.985
0.950
0.981
0.997
0.978
0.973
0.957
0.986
0.963
EC50 (µM)
268.58
220.54
14.73
3.23
4a
Y = 4.151X − 16.812
Y = 2.181X − 7.968
Y = 2.236X − 9.418
Y = 0.902X − 3.382
Y = 1.143X − 4.821
Y = 2.993X − 12.458
Y = 2.136X − 8.651
Y = 3.052X − 12.407
Y = 2.152X − 8.824
Y = 2.170X − 8.084
Y = 1.797X − 6.847
Y = 8.084X − 36.222
Y = 3.608X − 15.344
Y = 2.80X − 12.685
Y = 1.440X − 5.298
Y = 3.995X − 17.315
Y = 2.538X − 11.106
Y = 1.283X − 5.263
Y = 1.459X − 6.242
Y = 1.263X − 4.926
Y = 0.802X − 3.709
Y = 2.127X − 8.363
Y = 1.514X − 5.267
Y = 1.329X − 4.712
4e
4g
5g
Carbendazim
4.12
2a
120.78
38.25
218.48
24.18
149.00
48.73
4425.75
146.93
6.07
3a
R. solani
4a
R. solani
4d
R. solani
4g
R. solani
4i
R. solani
4j
R. solani
4l
R. solani
4m
R. solani
4n
R. solani
21.68
122.76
10.38
4.14
4o
R. solani
4t
R. solani
4e
A. solani
4i
A. solani
3.27
6
Fenamidone
4d
A. solani
9.86
A. solani
0.90
B. cinerea
B. cinerea
B. cinerea
52.49
51.02
38.92
4g
Fenamidone

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Figure 1. Inhibition of compound 4g against spore germination of B. cinerea. The pictures
were taken from the fungi growing in the glass plates with a microscope with Scopephoto
software (version 3.6, Photoscope Inc., Arlington, VA, USA, 2009) and magnified by
10 × 10 times without stain. The fungi were grown in clear water (CK) or sample solution
plus water (treated) at 25 ± 0.5 °C for 24 h. Color is from background.
3. Experimental Section
3.1. General Information
All reactions were performed with magnetic stirring. Unless otherwise stated, all reagents were
purchased from commercial suppliers and used without further purification. Organic solutions were
concentrated under reduced pressure using a rotary evaporator. Melting points were measured on a
1
Yanagimoto apparatus (Yanagimoto MFG Co., Kyoto, Japan) and are uncorrected. H-NMR spectra
were obtained on Bruker DPX 300 spectrometer (Bruker Biospin Co., Stuttgart, Germany) with CDCl3
or DMSO-d6 as a solvent and TMS as an internal standard. High resolution mass spectral analysis was
performed on a LTQ Orbitrap instrument (ThermoFisher scientific Inc., Waltham, MA, USA).
3.2. Synthesis
3.2.1. Synthesis of 3-Acetyl-4-methyl(phenyl)-5,5-dimethylbutenolide (1a and 1b)
The synthesis of the intermediates 1a and 1b were carried out according to the literature protocols
and their spectral data were identical with those in the references [18,20]. Compound 1a, colorless
solid, yield 80%, m.p. 50–51 °C, 1H-NMR (CDCl3, 300 MHz) δ: 2.47 (s, 3H), 2.28 (s, 3H), 1.42 (s, 6H).
1
Compound 1b, colorless solid, yield 84%, m.p. 76–77 °C, H-NMR (CDCl3, 300 MHz) δ: 7.48–7.46
(m, 3H), 7.25–7.22 (m, 2H), 2.39 (s, 3H), 1.56 (s, 3H).
3.2.2. Synthesis of 4-Methyl(phenyl)-5,5-dimethyl-3-ethylidenebutenolide-5-thiohydantoin (2a and 2b)
The synthesis of the intermediates 2a and 2b were performed following the processes in our
previous paper [10,11] by reaction of 1a and 1b with thiohydantoin. Compound 2a, yellow solid, yield
1
63%, m.p. 218–220 °C, H-NMR (DMSO-d6, 300 MHz) δ: 12.12 (s, 1H), 11.99 (s, 1H), 2.03 (s, 3H),
1.87 (s, 3H), 1.42 (s, 6H); HR-ESI-MS m/z: calcd for C12H15N2O3S [M + H]+ 267.0798; found,
267.0798. Compound 2b, yellow solid, yield 67%, m.p. 223–225 °C, 1H-NMR (DMSO-d6, 300 MHz)

Molecules 2015, 20
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δ: 12.20 (s, 1H), 11.99 (s,1H), 7.51–7.45 (m, 3H), 7.35–7.30 (m, 2H), 1.80 (s, 3H), 1.63 (s, 3H), 1.46
(s, 3H); HR-ESI-MS m/z: calcd for C17H17N2O3S [M + H]+ 329.0954; found, 329.0944.
3.2.3. Synthesis of 4-Methyl(phenyl)-5,5-dimethyl-3-ethylidenebutenolide-5-(2-methylthioimidazo-4-one)
(3a and 3b)
The synthesis of the intermediates 3a and 3b were performed according to the methods in our
previous paper [10,11] by reaction of 2a and 2b with methyl iodide. Compound 3a, yellow solid, yield
1
90%, m.p. 168–170 °C, H-NMR (CDCl3, 300 MHz) δ: 10.23 (s, 1H), 2.61 (s, 3H), 2.37(s, 3H), 1.90
(s, 3H), 1.54 (s, 3H), 1.50 (s, 3H); HR-ESI-MS m/z: calcd for C13H17N2O3S [M + H]+ 281.0954; found,
1
281.0955. Compound 3b, yellow solid, yield 79%, m.p. 204–206 °C, H-NMR (CDCl3, 300 MHz)
δ: 10.16 (s, 1H), 7.42–7.37 (m, 3H), 7.27–7.23 (m, 2H), 2.57 (s, 3H), 2.08 (s, 3H), 1.68 (s, 3H), 1.59
(s, 3H); HR-ESI-MS m/z: calcd for C17H17N2O3S [M + H]+ 343.1111; found, 343.1102.
3.2.4. General Procedure for the Synthesis of Compounds 4 and 5
To a stirred solution of 1.0 mmol 3a, or 3b in 20 mL of acetic acid or oxalic acid and dioxane, the
amines were added at ambient temperature and heated to reflux for 10–24 h. The reactions were monitored
by TLC. After completion, the solvents were removed under reduced pressure. The residues were purified
by silica gel chromatography using CH2Cl2/acetone as eluents to afford compounds 4 and 5.
5-(4,5,5-Trimethyl-3-ethylidenebutenolide)-2-phenylaminoimidazolinone 4a, white solid, yield 65%,
1
m.p. 203–206 °C, H-NMR (DMSO-d6, 300 MHz) δ: 10.44–9.68 (m, 2H), 7.71–6.99 (m, 5H), 2.12
(s, 3H), 1.86 (s, 3H), 1.43 (s, 6H); HR-ESI-MS m/z: calcd for C18H20N3O3 [M + H]+ 326.1499;
found, 326.1499.
5-(4,5,5-Trimethyl-3-ethylidenebutenolide)-2-(4-methylphenyl)aminoimidazolinone 4b, white solid,
yield 83%, m.p. 208–210 °C, 1H-NMR (DMSO-d6, 300 MHz) δ: 10.40–9.52 (m, 2H), 7.59–7.12 (m, 4H),
2.26 (s, 3H), 2.09 (s, 3H), 1.85 (s, 3H), 1.43 (s, 6H); HR-ESI-MS m/z: calcd for C19H22N3O3 [M + H]+
340.1656; found, 340.1655.
5-(4,5,5-Trimethyl-3-ethylidenebutenolide)-2-(4-flurophenyl)aminoimidazolinone 4c, white solid, yield
56%, m.p. 194–196 °C, 1H-NMR (DMSO-d6, 300 MHz) δ: 10.53–9.66 (m, 2H), 7.71–7.14 (m, 4H), 2.10
(s, 3H), 1.85 (s, 3H), 1.43 (s, 6H); HR-ESI-MS m/z: calcd for C18H19FN3O3 [M + H]+ 344.1405;
found, 344.1405.
5-(4,5,5-Trimethyl-3-ethylidenebutenolide)-2-(4-methoxyphenyl)aminoimidazolinone 4d, yellow solid,
yield 87%, m.p. 151–154 °C, 1H-NMR (DMSO-d6, 300 MHz) δ: 10.60–9.61 (m, 2H), 7.54 (d, J = 7.5 Hz,
2H), 6.88 (d, J = 7.5 Hz, 2H), 3.74 (s, 3H), 2.31 (s, 3H), 1.91 (s, 3H), 1.44 (s, 6H); HR-ESI-MS m/z:
calcd for C19H22N3O4 [M + H]+ 356.1605; found, 356.1605.
5-(4,5,5-Trimethyl-3-ethylidenebutenolide)-2-(4-trifluromethylphenyl)aminoimidazolinone 4e, yellow
solid, yield 59%, m.p. 203–206 °C, 1H-NMR (DMSO-d6, 300 MHz) δ: 10.73–10.06 (m, 2H), 8.00–7.66

Molecules 2015, 20
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(m, 4H), 2.12 (s, 3H), 1.86 (s, 3H), 1.43 (s, 6H); HR-ESI-MS m/z: calcd for C19H19F3N3O3 [M + H]+
394.1373; found, 394.1374.
N-Methyl-5-(4,5,5-trimethyl-3-ethylidenebutenolide)-2-phenylaminoimidazolinone 4f, white solid,
yield 56%, m.p. 195–198 °C, 1H-NMR (DMSO-d6, 300 MHz) δ: 7.54–7.44 (m, 5H), 3.61 (s, 3H), 2.32
(s, 3H), 1.90 (s, 3H), 1.41 (s, 6H); HR-ESI-MS m/z: calcd for C19H22N3O3 [M + H]+ 340.1656;
found, 340.1655.
5-(4,5,5-Trimethyl-3-ethylidenebutenolide)-2-(3-trifluromethylphenyl)aminoimidazolinone 4g, yellow
solid, yield 66%, m.p. 148–151 °C, 1H-NMR (DMSO-d6, 300 MHz) δ: 10.70–9.82 (m, 2H), 8.20–7.30
(m, 4H), 2.33 (s, 3H), 1.86 (s, 3H), 1.47 (s, 6H); HR-ESI-MS m/z: calcd for C19H19F3N3O3 [M + H]+
394.1373; found, 394.1374.
5-(4,5,5-Trimethyl-3-ethylidenebutenolide)-2-(2-methylphenyl)aminoimidazolinone 4h, white solid,
yield 89%, m.p. 184–187 °C, 1H-NMR (DMSO-d6, 300 MHz) δ: 10.38–9.00 (m, 2H), 7.20–6.97 (m, 4H),
2.26 (s, 3H), 2.09 (s, 3H), 1.84 (s, 3H), 1.42 (s, 6H); HR-ESI-MS m/z: calcd for C19H22N3O3 [M + H]+
340.1656; found, 340.1654.
5-(4,5,5-Trimethyl-3-ethylidenebutenolide)-2-(2-flurophenyl)aminoimidazolinone 4i, white solid, yield
74%, m.p. 166–169 °C, 1H-NMR (DMSO-d6, 300 MHz) δ: 10.54–9.87 (m, 2H), 7.68–7.21 (m, 4H), 2.32
(s, 3H), 2.00 (s, 3H), 1.48 (s, 6H); HR-ESI-MS m/z: calcd for C18H19FN3O3 [M + H]+ 344.1405;
found, 344.1406.
5-(4,5,5-Trimethyl-3-ethylidenebutenolide)-2-(2-methoxyphenyl)aminoimidazolinone 4j, yellow solid,
yield 58%, m.p. 154–156 °C, 1H-NMR (DMSO-d6, 300 MHz) δ: 10.41–9.65 (m, 2H), 7.07–6.90 (m, 4H),
3.88 (s, 3H), 2.09 (s, 3H), 1.92 (s, 3H), 1.50 (s, 6H); HR-ESI-MS m/z: calcd for C19H22N3O4 [M + H]+
356.1605; found, 356.1606.
5-(4,5,5-Trimethyl-3-ethylidenebutenolide)-2-(2-chlororophenyl)aminoimidazolinone 4k, white solid,
1
yield 23%, m.p.165–168 °C, H-NMR (DMSO-d6, 300 MHz) δ: 10.41–9.92 (m, 2H), 7.44–7.00 (m,
4H), 2.09 (s, 3H), 1.85 (s, 3H), 1.41 (s, 6H); HR-ESI-MS m/z: calcd for C18H19ClN3O3 [M + H]+
360.1110; found, 360.1110.
5-(4,5,5-Trimethyl-3-ethylidenebutenolide)-2-(2,6-dimethylphenyl)aminoimidazolinone 4l, white solid,
yield 76%, m.p. 240–243 °C, 1H-NMR (DMSO-d6, 300 MHz) δ: 10.41–9.19 (m, 2H), 7.12–6.95 (m, 3H),
2.11 (s, 6H), 1.84 (s, 3H), 1.44 (s, 6H); HR-ESI-MS m/z: calcd for C19H22N3O3 [M + H]+ 340.1656;
found, 340.1654.
5-(4,5,5-Trimethyl-3-ethylidenebutenolide)-2-benzylaminoimidazolinone 4m, white solid, yield 30%,
1
m.p. 168–170 °C, H-NMR (DMSO-d6, 300 MHz) δ: 10.57–9.81 (m, 2H), 7.34–7.25 (m, 5H), 4.51
(brs, 2H), 1.99 (s, 3H), 1.80 (s, 3H), 1.39 (s, 6H); HR-ESI-MS m/z: calcd for C19H22N3O3 [M + H]+
340.1656; found, 340.1655.

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5-(4,5,5-Trimethyl-3-ethylidenebutenolide)-2-(4-methoxybenzyl)aminoimidazolinone 4n, white solid,
yield 56%, m.p. 145–148 °C, 1H-NMR (DMSO-d6, 300 MHz) δ: 10.57–9.81 (m, 2H), 7.34–7.25 (m, 5H),
4.51 (brs, 2H), 3.73 (s, 3H), 1.99 (s, 3H), 1.80 (s, 3H), 1.39 (s, 6H); HR-ESI-MS m/z: calcd for
C20H24N3O4 [M + H]+ 370.1761; found, 370.1761.
5-(4,5,5-Trimethyl-3-ethylidenebutenolide)-2-(4-chlrobenzyl)aminoimidazolinone 4o, white solid, yield
54%, m.p. 147–150 °C, 1H-NMR (DMSO-d6, 300 MHz) δ: 10.79–9.07 (m, 2H), 7.43–7.34 (m, 4H), 4.48
(brs, 2H), 1.98 (s, 3H), 1.81 (s, 3H), 1.40 (s, 6H); HR-ESI-MS m/z: calcd for C19H21ClN3O3 [M + H]+
374.1266; found, 374.1267.
5-(4,5,5-Trimethyl-3-ethylidenebutenolide)-2-(4-flurobenzyl)aminoimidazolinone 4p, yellow solid,
yield 59%, m.p. 140–143 °C, 1H-NMR (DMSO-d6, 300 MHz) δ: 10.57–8.32 (m, 2H), 7.62–6.77 (m, 4H),
4.47 (brs, 2H), 1.99 (s, 3H), 1.81 (s, 3H), 1.40 (s, 6H); HR-ESI-MS m/z: calcd for C19H21FN3O3 [M + H]+
358.1562; found, 358.1561.
5-(4,5,5-Trimethyl-3-ethylidenebutenolide)-2-(3-trifluromethylbenzyl)aminoimidazolinone 4q, yellow
solid, yield 48%, m.p. 127–130 °C, 1H-NMR (DMSO-d6, 300 MHz) δ: 10.84–9.15 (m, 2H), 7.72–7.48
(m, 4H), 4.58 (brs, 2H), 1.97 (s, 3H), 1.81 (s, 3H), 1.44 (s, 6H); HR-ESI-MS m/z: calcd for
C20H21F3N3O3 [M + H]+ 408.1530; found, 408.1528.
5-(4,5,5-Trimethyl-3-ethylidenebutenolide)-2-(2-flurobenzyl)aminoimidazolinone 4r, yellow solid,
yield 81%, m.p. 227–230 °C, 1H-NMR (DMSO-d6, 300 MHz) δ: 10.51–9.84 (m, 2H), 7.45–7.18 (m, 4H),
4.52 (brs, 2H), 1.99 (s, 3H), 1.82 (s, 3H), 1.41 (s, 6H); HR-ESI-MS m/z: calcd for C19H21FN3O3 [M + H]+
358.1562; found, 358.1562.
5-(4,5,5-Trimethyl-3-ethylidenebutenolide)-2-(2-chlorobenzyl)aminoimidazolinone 4s, white solid,
yield 70%, m.p. 164–166 °C, 1H-NMR (DMSO-d6, 300 MHz) δ: 10.41–9.92 (m, 2H), 7.53–7.38 (m, 4H),
4.71 (brs, 2H), 2.32 (s, 3H), 2.01 (s, 3H), 1.44 (s, 6H); HR-ESI-MS m/z: calcd for C19H21ClN3O3 [M + H]+
374.1266; found, 374.1267.
5-(4,5,5-Trimethyl-3-ethylidenebutenolide)-2-(2-methoxybenzyl)aminoimidazolinone 4t, yellow solid,
yield 34%, m.p. 120–123 °C, 1H-NMR (DMSO-d6, 300 MHz) δ: 10.40–8.83 (m, 2H), 7.30–6.91 (m, 4H),
4.46 (brs, 2H), 3.83 (s, 3H), 1.96 (s, 3H), 1.81 (s, 3H), 1.44 (s, 6H); HR-ESI-MS m/z: calcd for
C20H24N3O4 [M + H]+ 370.1761; found, 370.1758.
5-(4-Phenyl-5,5-dimethyl-3-ethylidenebutenolide)-2-phenylaminoimidazolinone 5a, yellow solid, yield
44%, m.p. 167–170 °C, 1H-NMR (DMSO-d6, 300 MHz) δ: 10.52–9.34 (m, 2H), 7.68–6.99 (m, 10H), 1.88
(s, 3H), 1.69 (s, 3H), 1.48 (s, 3H); HR-ESI-MS m/z: calcd for C23H22N3O3 [M + H]+ 388.1656;
found, 388.1644.
5-(4-Phenyl-5,5-dimethyl-3-ethylidenebutenolide)-2-(4-methylphenyl)aminoimidazolinone 5b, yellow
solid, yield 60%, m.p. 185–188 °C, 1H-NMR (DMSO-d6, 300 MHz) δ: 10.47–9.32 (m, 2H), 7.47–7.04
(m, 9H), 2.25 (s, 3H), 1.87 (s, 3H), 1.61 (s, 3H), 1.47 (s, 3H); HR-ESI-MS m/z: calcd for C24H24N3O3
[M + H]+ 402.1812; found, 402.1801.

Molecules 2015, 20
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5-(4-Phenyl-5,5-dimethyl-3-ethylidenebutenolide)-2-(4-flurophenyl)aminoimidazolinone 5c, yellow
solid, yield 47%, m.p. 164–167 °C, 1H-NMR (DMSO-d6, 300 MHz) δ: 10.61–9.37 (m, 2H), 7.71–7.08
(m, 9H), 1.87 (s, 3H), 1.61 (s, 3H), 1.48 (s, 3H); HR-ESI-MS m/z: calcd for C23H21FN3O3 [M + H]+
406.1561; found, 406.1549.
5-(4-Phenyl-5,5-dimethyl-3-ethylidenebutenolide)-2-(4-methoxyphenyl)aminoimidazolinone 5d, yellow
solid, yield 82%, m.p. 217–219 °C, 1H-NMR (DMSO-d6, 300 MHz) δ: 10.44–9.21 (m, 2H), 7.58–6.85
(m, 9H), 3.74 (s, 3H), 1.85 (s, 3H), 1.61 (s, 3H), 1.47 (s, 3H); HR-ESI-MS m/z: calcd for C24H24N3O4
[M + H]+ 418.1761; found, 418.1749.
5-(4-Phenyl-5,5-dimethyl-3-ethylidenebutenolide)-2-(4-trifluromethylphenyl)aminoimidazolinone 5e,
1
white solid, yield 86%, m.p. 190–193 °C, H-NMR (DMSO-d6, 300 MHz) δ: 10.45–9.35 (m, 2H),
7.97–7.30 (m, 9H), 1.89 (s, 3H), 1.62 (s, 3H), 1.48 (s, 3H); HR-ESI-MS m/z: calcd for C24H21F3N3O3
[M + H]+ 456.1530; found, 456.1518.
5-(4-Phenyl-5,5-dimethyl-3-ethylidenebutenolide)-2-(3-trifluromethylphenyl)aminoimidazolinone 5f,
1
white solid, yield 77%, m.p. 169–172 °C, H-NMR (DMSO-d6, 300 MHz) δ: 10.77–9.57 (m, 2H),
8.23–7.32 (m, 9H), 1.90 (s, 3H), 1.59 (s, 3H), 1.49 (s, 3H); HR-ESI-MS m/z: calcd for C24H21F3N3O3
[M + H]+ 456.1530; found, 456.1517.
5-(4-Phenyl-5,5-dimethyl-3-ethylidenebutenolide)-2-(2-flurophenyl)aminoimidazolinone 5g, yellow
solid, yield 32%, m.p. 158–161 °C, 1H-NMR (DMSO-d6, 300 MHz) δ: 10.51–9.22 (m, 2H), 7.62–7.04
(m, 9H), 1.80 (s, 3H), 1.61 (s, 3H), 1.45 (s, 3H); HR-ESI-MS m/z: calcd for C23H21FN3O3 [M + H]+
406.1561; found, 406.1549.
5-(4-Phenyl-5,5-dimethyl-3-ethylidenebutenolide)-2-(2-chlorophenyl)aminoimidazolinone 5h, yellow
solid, yield 32%, m.p. 158–161 °C, 1H-NMR (DMSO-d6, 300 MHz) δ: 10.73–9.48 (m, 2H), 7.50–7.00
(m, 9H), 1.91 (s, 3H), 1.60 (s, 3H), 1.44 (s, 3H); HR-ESI-MS m/z: calcd for C23H21ClN3O3 [M + H]+
406.1561; found, 406.1549.
5-(4-Phenyl-5,5-dimethyl-3-ethylidenebutenolide)-2-(2-methoxyphenyl)aminoimidazolinone 5i, yellow
solid, yield 84%, m.p. 236–239 °C, 1H-NMR (DMSO-d6, 300 MHz) δ: 10.18–9.41 (m, 2H), 7.46–6.94
(m, 9H), 3.88 (s, 3H), 1.89 (s, 3H), 1.61 (s, 3H), 1.47 (s, 3H); HR-ESI-MS m/z: calcd for C24H24N3O4
[M + H]+ 418.1761; found, 418.1748.
5-(4-Phenyl-5,5-dimethyl-3-ethylidenebutenolide)-2-benzylaminoimidazolinone 5j, white solid, yield
1
55%, m.p. 128–131 °C, H-NMR (DMSO-d6, 300 MHz) δ: 10.60–9.33 (m, 2H), 7.45–7.27 (m, 10H),
4.51 (brs, 2H), 1.80 (s, 3H), 1.59 (s, 3H), 1.44 (s, 3H); HR-ESI-MS m/z: calcd for C24H24N3O3
[M + H]+ 402.1812; found, 402.1801.
5-(4-Phenyl-5,5-dimethyl-3-ethylidenebutenolide)-2-(4-chlorobenzyl)aminoimidazolinone 5k, yellow
solid, yield 67%, m.p. 140–143 °C, 1H-NMR (DMSO-d6, 300 MHz) δ: 10.67–9.40 (m, 2H), 7.46–7.17
(m, 9H), 4.46 (brs, 2H), 1.74 (s, 3H), 1.59 (s, 3H), 1.44 (s, 3H); HR-ESI-MS m/z: calcd for
C24H23ClN3O3 [M + H]+ 436.1422; found, 436.1412.

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5-(4-Phenyl-5,5-dimethyl-3-ethylidenebutenolide)-2-(2-methoxybenzyl)aminoimidazolinone 5l, yellow
solid, yield 67%, m.p. 116–119 °C, 1H-NMR (DMSO-d6, 300 MHz) δ: 10.18–9.41 (m, 2H), 7.46–6.94
(m, 9H), 3.88 (s, 3H), 1.89 (s, 3H), 1.59 (s, 3H), 1.44 (s, 3H); HR-ESI-MS m/z: calcd for C25H26N3O4
[M + H]+ 432.1918; found, 432.1916.
4,5,5-Trimethyl-3-ethylidenebutenolide-5-hydantoin 6, white solid, yields 10%–40% isolated from the
1
mixtures of preparing reactions 4a to 4t as byproducts, m.p. 230–232 °C, H-NMR (DMSO-d6,
300 MHz) δ: 10.98 (s, 1H), 10.26 (s, 1H), 1.90 (s, 3H), 1.85 (s, 3H), 1.41 (s, 6H); 13C-NMR (DMSO-d6,
75 MHz) δ: 170.02, 167.04, 162.88, 154.54, 19.21, 124.00, 117.70, 85.83, 24.19, 24.09, 17.93, 11.73;
HR-ESI-MS m/z: calcd for C12H15N2O4 [M + H]+ 251.1026; found, 251.1026.
3.3. Bioassay of Fungicidal Activity
The preliminary fungicidal activities of compounds 4–6 against S. scleotiorum, R. solani, P. capsici,
A. solani, B. cinerea and F. graminearum were evaluated using methods in the references [21–23] by the
mycelium growth rate and spore germination tests [24,25]. The cultures were incubated at 25 ± 0.5 °C.
Three replicates were performed and the mean measurements were calculated from the three replicates.
The EC50 values were determined from the inhibition rates of five different concentrations based on the
statistics method for the compounds which had more than 65% inhibition rates. Chlorothalonil,
carbendazim and fenamidone were used as positive controls in the mycelium growth rate test, while
pure water was used as blank control in the spore germination test.
4. Conclusions
The novel 5-(5,5-dimethylbutenolide-3-ethylidene)-2-aminoimidazolinone derivatives were designed
and synthesized in moderate to excellent yields in four steps by α-hydroxyketone and diketene as starting
1
materials. The products were characterized by HR-ESI-MS and H-NMR. The preliminary bioassay
showed that some of these compounds, such as 4a, 4e and 5g exhibit 94.9%, 92.8% and 81.4%
inhibition rates against S. scleotiorum, respectively. The EC50 values of compounds 4e and 4i were
4.14 and 3.27 µM against A. Solani, while 5g had EC50 value of 3.23 µM against S. scleotiorum.
Compounds 4d and 4g had 98.0% and 97.8% inhibition rates of spore germination against B. cinerea
at the concentration of 100 µg/mL, respectively. Further syntheses and structure optimization studies
are in progress in our laboratory.
Supplementary Materials
Supplementary materials can be accessed at: http://www.mdpi.com/1420-3049/20/08/13740/s1.
Acknowledgments
This project was co-funded by the program of Ministry of Education, China (No. 20130008110003),
National Natural Science Foundation of China (No. 21172254) and the 12th Five-year National Key
Technologies R & D Program of China (No. 2011BAE06B03).

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Author Contributions
B. Tang, M. Yang and Y. Zhao synthesized all of novel compounds, L. Kong and W. Wang run the
bioassay evaluation and statistics analysis, M. Wang started the project, designed the molecules and
wrote the paper.
Conflict of Interest
The authors declare no conflict of interest.
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Sample Availability: Samples of the compounds 4a–4t, 5a–5l and 6 are available from the authors.
© 2015 by the authors; licensee MDPI, Basel, Switzerland. This article is an open access article
distributed under the terms and conditions of the Creative Commons Attribution license
(http://creativecommons.org/licenses/by/4.0/).