"amano" lipase DF-catalyzed efficient synthesis of 2,2′-arylmethylene dicyclohexane-1,3-dione derivatives in anhydrous media

Article abstract of DOI:10.1016/j.cclet.2014.04.007

A simple and efficient method was developed for the synthesis of 2,2′-arylmethylene dicyclohexane-1,3-dione derivatives via the Knoevenagel-Michael cascade reactions of aromatic aldehydes and 1,3-cyclic diketones catalyzed by "Amano" lipase DF, which expands the application field of enzyme catalytic promiscuity. This protocol provides several advantages over the traditional chemical synthesis, such as simple work-up procedure, high yields (up to 94%) and environmental friendliness.

Full text of DOI:10.1016/j.cclet.2014.04.007

G Model
CCLET 2944 1–3
Chinese Chemical Letters xxx (2014) xxx–xxx
Contents lists available at ScienceDirect
Chinese Chemical Letters
journal homepage: www.elsevier.com/locate/cclet
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Original article
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‘‘Amano’’ lipase DF-catalyzed efficient synthesis of 2,20-arylmethylene
dicyclohexane-1,3-dione derivatives in anhydrous media
Q1 Ling Jiang a, Bo Wang b, Rong-Rong Li c, Sa Shen c, Hong-Wei Yu a, Li-Dan Ye a,
*
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a Department of Chemical and Biological Engineering, Zhejiang University, Hangzhou 310007, China
b College of Materials and Chemical Engineering, Hainan University, Haikou 570228, China
c School of Chemical and Material Engineering, Jiangnan University, Wuxi 214122, China
A R T I C L E I N F O
A B S T R A C T
A simple and efficient method was developed for the synthesis of 2,20-arylmethylene dicyclohexane-1,3-
dione derivatives via the Knoevenagel–Michael cascade reactions of aromatic aldehydes and 1,3-cyclic
diketones catalyzed by ‘‘Amano’’ lipase DF, which expands the application field of enzyme catalytic
promiscuity. This protocol provides several advantages over the traditional chemical synthesis, such as
simple work-up procedure, high yields (up to 94%) and environmental friendliness.
Article history:
Received 30 December 2013
Received in revised form 6 March 2014
Accepted 20 March 2014
Available online xxx
ß 2014 Li-Dan Ye. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved.
Keywords:
‘‘Amano’’ lipase DF
Knoevenagel condensation
Michael addition
Catalytic promiscuity
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1. Introduction
lower energy requirements [5]. One current frontier for biocataly- 32
sis is enzymatic promiscuity, where enzymes catalyze alternative 33
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Derivatives of 2,20-arylmethylene dicyclohexane-1,3-dione,
which serve as key intermediates in the synthesis of xanthenes,
an important class of biological active compounds, are typically
synthesized via the Knoevenagel condensation and Michael
addition of aromatic aldehydes with 1,3-cyclohexanedione, or
other types of 1,3-cyclic diketones. Jin et al. carried out the
condensation by grinding at room temperature [1]. Kantevari and
coworkers used HClO4–SiO2 as catalyst to synthesize the title
compounds in 44.5–91.0% yields [2]. In the presence of CsF, Nandre
et al. synthesized the target product in 84–96% yields [3]. Recently,
Li et al. reported that the condensation can be catalyzed by urea
under ultrasound [4]. However, these reported chemical synthetic
methods suffer from a number of drawbacks, such as low yield,
harsh reaction conditions and tedious work-up processes. To fulfill
the requirements of green chemistry and to simplify the
production process, a simple, efficient and eco-friendly method
should be developed for the synthesis of 2,20-arylmethylene
dicyclohexane-1,3-dione derivatives.
reactions that differ from their natural physiological reaction. The 34
need for new efficient and eco-friendly catalysts in industry is 35
evident and enzymatic promiscuity can meet these requirements 36
well, which has largely broadened the applicable range of 37
biocatalysis in the synthesis of fine chemicals [6]. Examples of 38
enzyme promiscuity-related reactions include Aldol reaction [7], 39
Baylis–Hillman reaction [8] and Mannich reaction [9], while no 40
enzyme catalyzed Knoevenagel–Michael cascade reactions of 41
aromatic aldehydes and 1,3-cyclic diketones have been reported. 42
Among the promiscuous enzymes, lipases, which naturally 43
hydrolyze triglycerides, have attracted much attention due to 44
their excellent features, such as wide sources, broad substrate 45
spectrum and high stability. In continuation of our work [10] on 46
the enzymatic promiscuity, herein we report a novel method for 47
the synthesis of 2,20-arylmethylene dicyclohexane-1,3-dione 48
derivatives using ‘‘Amano’’ lipase DF as a catalyst (Scheme 1).
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2. Experimental
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During the last decade, biocatalysis has emerged as an efficient
and powerful tool in modern synthetic chemistry due to its valued
features, such as high efficiency, mild reaction conditions and
Analytical thin layer chromatography (TLC) was performed on 51
Haiyang precoated TLC plates (silica gel GF254), using petroleum 52
ether/ethyl acetate (1/1, v/v) as the elution solution. The 1H NMR 53
(400 MHz) spectra were recorded on a Bruker Advance 2B 400 54
instrument. Chemical shifts (
NMR 77.16) as the solvent and tetramethylsilane 56
7.26, 13C NMR
d
) are quoted in ppm using CDCl3 (1H 55
*
Corresponding author.
E-mail address: yelidan@zju.edu.cn (L.-D. Ye).
d
d
http://dx.doi.org/10.1016/j.cclet.2014.04.007
1001-8417/ß 2014 Li-Dan Ye. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved.
Please cite this article in press as: L. Jiang, et al., ‘‘Amano’’ lipase DF-catalyzed efficient synthesis of 2,20-arylmethylene dicyclohexane-
1,3-dione derivatives in anhydrous media, Chin. Chem. Lett. (2014), http://dx.doi.org/10.1016/j.cclet.2014.04.007

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CCLET 2944 1–3
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L. Jiang et al. / Chinese Chemical Letters xxx (2014) xxx–xxx
Table 1
Optimization of the reaction conditions for the synthesis of 3a.a
Entry Catalyst
Enzyme concentration Yield (%)b
(mg/mL)
1
2
–
–
20
20
20
20
20
20
10
15
30
40
0
12
9
Scheme 1. ‘‘Amano’’ Lipase DF-catalyzed synthesis of 2,20-arylmethylene
‘‘Amano’’ lipase AK
‘‘Amano’’ lipase AS
‘‘Amano’’ lipase DF
CAL-B
dicyclohexane-1,3-dione derivatives.
3
4
89
21
18
29
58
75
91
90
3
57
58
59
60
61
62
63
64
65
66
67
68
69
70
71
72
73
74
75
76
77
78
79
80
81
82
83
84
85
86
87
(TMS) as the internal reference. All reagents were of analytical
grade and used as received without any further purification. HPLC
was carried out on a Agilent 1100 HPLC system equipped with a
Daicel 19423 CHIRALPAK AD-H column, using n-hexane/isopro-
panol (90:10, v/v) as the mobile phase at a flow rate of 1.0 mL/min
and UV detection at 254 nm. All melting points were determined
with a X-4 microscope melting-point testing apparatus and were
uncorrected. The structures of the products were confirmed by
comparing the NMR spectral data with those reported in the
literature.
5
6
Lipozyme RMIM
Lipozyme TLIM
‘‘Amano’’ lipase DF
‘‘Amano’’ lipase DF
‘‘Amano’’ lipase DF
‘‘Amano’’ lipase DF
7
8
9
10
11
12
Denatured ‘‘Amano’’ lipase DFc 20
a
Experimental conditions: a mixture of benzaldehyde (0.5 mmol), 1,3-cyclo-
hexanedione (0.5 mmol), enzyme and N,N-dimethylformamide (DMF) (5 mL) was
shaken at 200 rpm at 40 8C for 5 h.
b
Determined by HPLC using the external standard method.
General procedure for the synthesis of target compounds (3a–
3m): A mixture of aromatic aldehydes (0.5 mmol), 1,3-cyclic
diketone (0.5 mmol) and ‘‘Amano’’ lipase DF (20 mg/mL) in N,N-
dimethylformamide (DMF) (5 mL) was shaken at 200 rpm, 40 8C
for specified time. Progress of the reaction was monitored by TLC.
After completion of the reaction, 10 mL cold distilled water was
added to the reaction mixture, then extracted with dichloro-
methane (2ꢀ 15 mL). The organic phases were combined,
concentrated and purified by silica gel column chromatography
(petroleum ether:ethyl acetate = 2:1, v/v) or recrystallization from
ethanol to give pure product.
c
Pre-treated with urea at 100 8C for 8 h.
With the optimal reaction conditions in hand, a series of
aromatic aldehydes and 1,3-cyclic diketones were investigated to
test the generality and scope of this ‘‘Amano’’ lipase DF-catalyzed
Knoevenagel–Michael cascade reactions. The results are shown in
Table 2. Aromatic aldehydes bearing both electron-donating and
electron-withdrawing groups formed corresponding 2,20-aryl-
methylene dicyclohexane-1,3-dione derivatives in good to excel-
lent yields (Table 2, entries 1–13). Among the factors influencing
the reaction rate, the substituent position on the benzene ring of
aromatic aldehydes played an important role. The substituent in
the ortho-position led to a longer reaction time compared to that in
the para-position, which was most likely due to steric hinderance
(entry 12 vs. entry 13, Table 2). In addition, longer reaction times
were required with 5,5-dimethyl-1,3-cyclohexanedione in com-
parison to 1,3-cyclohexanedione, which can also be attributed to
steric hinderance (entries 1–5 vs. entries 6–13, Table 2).
114
115
116
117
118
119
120
121
122
123
124
125
126
127
128
129
130
131
132
133
134
135
136
137
Selected data of 3a: White solid; Mp 210–211 8C; 1H NMR
(400 MHz, CDCl3):
d 12.35 (s, 1H, OH), 12.19–11.88 (m, 1H, OH),
8.14–8.05 (m, 1H, Ar–H), 7.48 (t, 1H, J = 7.8 Hz, Ar–H), 7.28 (s, 0.3H,
Ar–H), 7.24 (s, 0.4H, Ar–H), 7.17 (t, 1H, J = 7.1 Hz, Ar–H), 7.10 (d, 1H,
J = 8.1 Hz, Ar–H), 5.47 (s, 1H, CH), 2.75–1.52 (m, 12H, CH2); 13C
NMR (101 MHz, CDCl3):
d 191.82, 189.79, 138.18, 133.70, 130.18,
128.50, 128.16, 126.48, 125.85, 116.45, 33.51, 32.99, 32.92, 20.13;
EI-MS [M]+m/z 312.
Characterization data of other compounds are shown in
Supporting information.
Based on these experimental results and the widely accepted
viewpoint that the hydrolysis active site in lipase is responsible for
its promiscuous performance [11], a mechanism is proposed and
depicted in Scheme 2. Firstly, 1,3-cyclic diketone was activated by
the Asp-His dyad to form an enolate anion, which was stabilized by
the oxyanion hole of the lipase. Then the enolate anion attacked the
aromatic aldehyde to form the intermediate I. Subsequently,
dehydration took place to generate the intermediate II. Finally,
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3. Results and discussion
89
90
91
92
93
94
95
96
Initially, benzaldehyde (1a) and 1,3-cyclohexanedione (2a)
were chosen as model substrates to optimize the reaction
conditions (Table 1). As expected, no product was observed in
the absence of the catalyst (Table 1, entry 1). In the cases of
‘‘Amano’’ lipase AK and ‘‘Amano’’ lipase AS, the target compound
3a was obtained in yields of only 12% and 9%, respectively (Table 1,
entries 2 and 3). CAL-B, Lipozyme RMIM and Lipozyme TLIM also
showed low catalytic activities in these cascade reactions, giving
the target product in yields of 21%, 18% and 29%, respectively
(entries 5–7, Table 1). When the reactants were incubated with
‘‘Amano’’ lipase DF, a yield of 89% was achieved, which was the
highest among all enzymes tested (Table 1, entry 4). As a control
experiment, the reactants were incubated with the urea-denatured
‘‘Amano’’ lipase DF, compound 3a was obtained only in 3% yield,
indicating the reaction was indeed catalyzed by the biologically
active ‘‘Amano’’ lipase DF (Table 1, entry 12).
Since enzyme concentration often plays an important role in
enzymatic reactions, concentrations of ‘‘Amano’’ lipase DF were
tested in the range of 10–40 mg/mL. As shown in Table 1, the yield
of the target product was evidently improved when the enzyme
concentration increased from 10 mg/mL to 20 mg/mL (Table 1,
entries 4, 8 and 9). However, further increase in the enzyme
concentration only led to slight change in the yield (Table 1, entries
10 and 11), suggesting that 20 mg/mL was a suitable concentration
for the catalysis of this cascade reactions.
Table 2
Synthesis of compound 3a–3m by lipase DF in DMF.
O
Ar
O
O
Lipase DF
R
R
R
R
R
R
97
98
99
Ar-CHO +
O
OH OH
Entry
Ar
R
Time (h)
Product
Yield (%)a
100
101
102
103
104
105
106
107
108
109
110
111
112
113
1
2
C6H5
H
5
5
5
5
5
6
8
6
8
6
6
8
6
3a
3b
3c
3d
3e
3f
89
91
90
84
88
83
85
92
81
87
86
93
94
4-CH3C6H4
4-CH3OC6H4
2-Furanyl
2-Thienyl
C6H5
H
3
H
4
H
5
H
6
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
7
2-FC6H4
3g
3h
3i
8
4-ClC6H4
9
3,4-Cl2C6H3
4-CH3C6H4
4-CH3OC6H4
2-O2NC6H4
4-O2NC6H4
10
11
12
13
3j
3k
3l
3m
a
Isolated yields.
Please cite this article in press as: L. Jiang, et al., ‘‘Amano’’ lipase DF-catalyzed efficient synthesis of 2,20-arylmethylene dicyclohexane-
1,3-dione derivatives in anhydrous media, Chin. Chem. Lett. (2014), http://dx.doi.org/10.1016/j.cclet.2014.04.007

G Model
CCLET 2944 1–3
L. Jiang et al. / Chinese Chemical Letters xxx (2014) xxx–xxx
3
Oxyanion hole
H
O
Oxyanion hole
H
His
His
His
O
Ar
O
O
O
Ar
H
-H2O
H
H
N
N
N
N
N
O
N
H
H
OH
H
H
H
H
H
O
H
O
O
O
O
O
H
Ar
O
O
O
I
Asp
Asp
Asp
O
O
Oxyanion hole
H
His
OH Ar
O
O
Ar
O
Ar
N
N
H
H
H
O
H
O
O
Ar
O
O
O
O
O
OH OH
III
Asp
II
O
O
O
Scheme 2. Proposed mechanism for ‘‘Amano’’ lipase DF-catalyzed synthesis of 2,20-arylmethylene dicyclohexane-1,3-dione derivatives.
138
139
Michael addition occurred when another enolate anion attacked
the intermediate II and gave the product III.
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140
4. Conclusion
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In summary, we have developed a novel method for the
enzymatic synthesis of 2,20-arylmethylene dicyclohexane-1,3-
dione derivatives via Knoevenagel–Michael cascade reactions
catalyzed by ‘‘Amano’’ lipase DF in anhydrous media. High yields,
mild reaction conditions, simple work-up procedure and eco-
friendliness are the features of this new protocol with great
potential in industrial applications.
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Acknowledgments
This work was financially supported by the Outstanding Young
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Natural Science Foundation of China (No. 21176215/21176102)
and the Program for Zhejiang leading team of S&T Innovation (No.
2011R50007). We thank all of the constructive comments and
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Appendix A. Supplementary data
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Supplementary data associated with this article can be found, in
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Please cite this article in press as: L. Jiang, et al., ‘‘Amano’’ lipase DF-catalyzed efficient synthesis of 2,20-arylmethylene dicyclohexane-
1,3-dione derivatives in anhydrous media, Chin. Chem. Lett. (2014), http://dx.doi.org/10.1016/j.cclet.2014.04.007