Cocatalytic enzyme system for the Michael addition reaction of in-situ-generated ortho-quinones

Article abstract of DOI:10.1002/ejoc.200800791

A laccase-lipase cocatalytic system was used to catalyze the domino reaction between catechols and nucleophilic reagents including 1,3-dicarbonyl compounds and aromatic amines in aqueous medium at room temperature. Lipase acted as a catalyst for Michael a

Full text of DOI:10.1002/ejoc.200800791

FULL PAPER
DOI: 10.1002/ejoc.200800791
Cocatalytic Enzyme System for the Michael Addition Reaction of
in-situ-Generated ortho-Quinones
Suteera Witayakran[a] and Arthur J. Ragauskas*[a,b]
Keywords: Enzyme catalysis / Laccase / Lipase / Michael addition / Cascade reaction / Quinones
A laccase-lipase cocatalytic system was used to catalyze the
domino reaction between catechols and nucleophilic rea-
gents including 1,3-dicarbonyl compounds and aromatic
amines in aqueous medium at room temperature. Lipase
acted as a catalyst for Michael addition step, and also en-
hanced the overall yield of the final products.
(© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim,
Germany, 2009)
by lipase. Although lipases (triacylglycerol hydrolase, EC
3.1.1.3) have been known to catalyze the hydrolysis and the
synthesis of esters formed from alcohols and acids,[7] recent
studies have reported the ability of lipases to catalyze
Michael addition reactions.[8] For example, Torre et al. pro-
vided the initial demonstration that lipase was able to cata-
lyze the Michael addition of secondary amines to acryloni-
trile.[8a] This reaction is clearly different from the natural
process this enzyme is usually associated with. Berglund et
al. has reported the Michael addition of 1,3-dicarbonyl
compounds to α,β-unsaturated carbonyl compounds cata-
lyzed by a C. antarctica lipase B mutant.[8b] Moreover,
Wang et al. recently established that lipase M from Mucor
javanicus is able to catalyze the Michael addition reaction
of pyrimidine with a disaccharide acrylate.[8c]
In a recent report, we demonstrated that laccase gener-
ated o-quinones could efficiently undergo Michael reactions
with 1,3-dicarbonyl compounds in the presence of
Sc(OTf)3 as a catalyst for the synthesis of benzofurans. On
average this reaction provided the adduct in 50–79% yield
but also produced a hazardous waste from the transitional
metal catalyst employed.[6l] From an environmental con-
cern, the development of alternative methodologies to re-
place the lanthanide metal catalyst in this synthesis is a high
priority to enhance the overall green chemistry aspect of
this one-pot synthetic reaction. Therefore, this report pres-
ents the use of the enzyme, lipase, as an alternative catalyst
for Michael addition reaction in conjugation with laccase
for the reaction of catechols with 1,3-dicarbonyl com-
pounds as well as with aromatic amines.
Introduction
In recent years, the advances in genomics and biotech-
nology have dramatically broadened the availability of low-
cost enzymes. In turn, this has increased the potential appli-
cation of enzymes for organic synthesis while also address-
ing the challenges of “green chemistry”.[1] A growing field
of interest in this field is the application of enzyme-initiated
domino reactions.[2] Under optimized reaction conditions,
it has been shown that several biocatalytic reactions can be
carried out in a single reactor.[3] For example, Kroutil and
his co-workers recently developed a one-pot, two-step, two-
enzyme cascade reaction for the synthesis of an enantiopure
epoxide.[4] Herein, we report on the use of two enzymes,
laccase and lipase, in the domino reaction of in-situ-gener-
ated o-quinones followed by enzyme-catalyzed Michael ad-
dition.
Laccase (benzenediol:oxygen oxidoreductase, EC
1.10.3.2) is a multinuclear-copper-containing oxidase that
has become an attractive biocatalyst that catalyzes the oxi-
dation of various aromatic compounds, including phenols,
o- and p-diphenols, and lignin derivatives.[5] This oxidore-
ductase enzyme frequently exhibits high oxidative selectiv-
ity in aqueous reactions and provides a unique green chem-
istry solution for organic synthesis.[6] In this study, laccase
was used to catalyze the oxidation of catechols to the corre-
sponding o-quinones which were reacted in-situ with nucle-
ophilic reagents, such as 1,3-dicarbonyl compounds and
aromatic amines, via a Michael addition reaction. In the
presence of lipase, the Michael addition step was catalyzed
[a] School of Chemistry and Biochemistry, Georgia Institute of
Technology,
Results and Discussion
Atlanta, GA 30332, USA
Fax: +1-404-8944778
E-mail: art.ragauskas@chemistry.gatech.edu
[b] Chalmers University of Technology, Forest Products and
Chemical Engineering Department, Chemical and Biological
Engineering,
1. Laccase-Lipase Cocatalytic System for the Reaction of
Catechols and 1,3-Dicarbonyl Compounds
In this study, laccase first catalyzed the oxidation of cate-
chols to the corresponding o-quinones which were reacted
Gothenburg, Sweden
Fax: +46-31-7722995
358
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Cocatalytic Enzyme System for Michael Addition Reaction
in-situ with 1,3-dicarbonyl compounds via a Michael ad- 2. In addition, lipase PS activity used in the reaction was
dition reaction. The Michael addition step was catalyzed much less than of lipase CR. Therefore, lipase PS was cho-
by lipase and the resulting addition product underwent a sen for further study. In order to verify whether the lipase
subsequent intramolecular cyclization to form benzofuran reaction is indeed catalyzed by the active site of lipase PS
derivative products (see Scheme 1).
and not by the protein, the reactions using inactivated li-
pase PS were conducted. The results in Table 1 show that
the inactivated lipase PS showed no catalytic activity for
these reactions.
Table 1. Reaction with a variety of lipases.
Scheme 1. Proposed reaction pathway of laccase/lipase catalytic
system for the synthesis of 3a.
In our initial studies, the reaction of catechol (1a) and
acetylacetone (2a) in the presence of laccase and lipase from
Candida rugosa (lipase CR, 60,000 U/mg) was investigated.
The reaction was carried out under atmospheric conditions
at room temperature (23 °C) in an aqueous buffered solu-
tion for 4 h. We found that the optimal yield of the product
(3a) of 60% was achieved when conducting the reaction of
1a and 2a in 1:2 molar ratio at pH 7.0, and using 100 U of
laccase and 10 mg (600,000 U) of lipase CR per 1 mmol of
1a. Because of the high activity of in situ-generated quin-
one, some side products (e.g. from the polymerization of
the quinone) were also observed but in this study we did
[a] Isolated yield.
To further define the catalytic benefits of lipase PS, the
not separate and identify these byproducts. For the control reaction of 1a and 2a in the presence of laccase with and
reaction when no laccase and lipase was added, no product without lipase PS were carried out. Sample aliquots were
3a was formed. When this reaction was preformed using taken every 30 min during the reaction and a quantitative
1
only lipase no product was formed, and in the presence of analysis of product 3a was measured by H NMR spec-
laccase only, the product 3a was formed in only 33% yield. troscopy using 1,3,5-trioxane as an internal standard. The
After this preliminary study, the next phase was to exam- calculated yield of the product 3a is higher than the isolated
ine a variety of lipases for this reaction system. These li- yield showed in Table 1 in both cases (with and without
pases included lipase CR (60,000 U/mg), lipase from Pseu- lipase PS). This can be explained by the lose of product
domonas cepacia (lipase PS, 46.2 U/mg), and lipase B Can- yield during the isolation process. However, in the end of
dida antarctica (CALB, 10.8 U/mg). The activity of these reaction, the yield difference between the reaction with and
lipases was measured by Aldrich methods which are dif- without lipase is about the same which is approximately
ferent for each lipase. The catalytic properties of these li- 25%. Figure 1 shows that in the beginning of the reaction,
pases were investigated by reacting 2a with catechol (1a) or the rate and yield of 3a from both reactions were almost
3-methylcatechol (1b), in the presence of laccase, as summa- the same. This can be explained by the predominance of
rized in Table 1. This study established that the optimal laccase-catalyzed oxidation of catechol at the beginning of
amount of each lipase to provide the highest yield of the the reaction and the gradually oxidation of catechol by lac-
product was different. The optimal amount of lipase CR, case which lead to a low concentration of o-quinone. How-
lipase PS and CALB for the reaction conditions used was ever, after 2 h of the reaction when the concentration of
600,000 U, 924 U, and 54 U per 1 mmol of catechol, respec- laccase-generated quinone was high the reaction with lipase
tively. The data in Table 1 shows that the yield of the prod- PS was predominant with less by-products and provided a
uct usually increased when lipase was added to the reaction. higher rate of the reaction and higher yield of the product
Lipase PS and lipase CR gave a high yield of the products than the reaction without lipase PS. Therefore, lipase PS
for both reactions while CALB was good only for reaction can enhance the overall yield for this reaction system.
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S. Witayakran, A. J. Ragauskas
FULL PAPER
catalytic effect on the reactions. In addition, the reactivity
of the 1,3-dicarbonyl compound employed also has an ef-
fect on efficiency of this two-enzyme system. When we used
1,3-dicarbonyl compounds that had an electron-with-
drawing group (Cl) at the alpha-position, the reaction was
complete in 1.5–2 h. The shorter reaction time was ascribed
to the increased acidity of the alpha-proton of these substi-
tuted 1,3-dicarbonyl compounds. The Cl atom was elimin-
ated during the reaction, and the proposed mechanism is
illustrated in Scheme 2. Besides 3-substituted catechols, 4-
substituted catechols, such as 4-chlorocatechol, can also be
used for the synthesis of polyhydroxylated benzofurans (en-
try 11). However, the yield of the product was low. In ad-
dition, we observed that this reaction system gave only one
isomer form of the possible benzofuran products.
Figure 1. The formation of compound 3a from the laccase-cata-
lyzed reaction of 1a and 2a in the presence and absence of lipase
PS. The percent yield of 3a was measured by 1H NMR spec-
troscopy.
Next, we examined the recyclability of this two-enzyme
catalytic system for the synthesis of benzofuran 3c. The
product 3c is relatively insoluble in the aqueous reaction
mixture and readily precipitates out of solution. Simple fil-
Following these optimization studies, we evaluated the tration of the product mixture facilitates reuse of the lipase/
breadth of this laccase-lipase cocatalytic system for the syn- laccase reaction system. The results shown in Table 3 dem-
thesis of benzofuran derivatives using a variety of catechols onstrate that this catalytic system can be reused for a sec-
and 1,3-dicarbonyl compounds. The results summarized in ond reaction, but for the third treatment only a low yield
Table 2 clearly suggest that the inactivated lipase has no of the product was formed.
Table 2. The reaction of catechols and 1,3-dicarbonyl compounds.
[a] Isolated yield. [b] Reaction time is 1.5–2 h.
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Cocatalytic Enzyme System for Michael Addition Reaction
result suggests that the increase of lipase dose did not pro-
vide a significant improvement for this reaction system
(Table 4). Characterization of the product by NMR and
mass spectrum indicated that the product was composed of
a 1:2 ratio of 1,2-benzoquinone and aniline (M+/z = 290).
Moreover, the product was not a quinone structure because
the carbonyl carbon signal was not observed in 13C-NMR
spectrum. The proposed reaction pathway for this reaction
and the structure of product (5a) was illustrated in
Scheme 2. Product 5a is known compound. Our NMR and
mass data are consistent with those in the literature.[11]
Scheme 2. Proposed mechanism pathway of the elimination of Cl
for the laccase/lipase-catalyzed reaction of catechol and chloro-1,3-
dicarbonyl compounds.
Table 3. Recycling of the catalytic system for the synthesis of 3-
acetyl-5,6-dihydroxy-2,7-dimethylbenzofuran (3c).
Run
Yield[a] (%) of 3c
1
2
3
72
62
5
Scheme 3. Proposed reaction pathway of laccase/lipase catalytic
system for the reaction between catechol (1a) and aniline (4a).
[a] Isolated Yield.
2. Laccase-Lipase Cocatalytic System for the Reaction of
Catechols and Anilines
Table 4. Reactions of catechol and anilines.
Next, we explored the feasibility of the laccase-lipase co-
catalytic system for the reaction between catechol and aro-
matic amines, anilines. Lalk and his co-worker have demon-
strated the ability to synthesize aminoquinones by laccase
initiated oxidation of p-hydroxyquinones followed by
Michael addition of primary aromatic amines in a good to
excellent yields.[6a] In contrast, herein, this nuclear anima-
tion reaction with the reactive 1,2-catechols was reported
to yield the corresponding products less than 35%. In the
presence of lipase, we had hypothesized that thus enzyme
could catalyze the Michael addition step of the reaction be-
tween the laccase-generated o-quinone and anilines thereby
improving the overall yields. We first conducted the reaction
of catechol (1a) (Scheme 3) and aniline (4a) in the presence
of laccase, with or without lipase PS, in phosphate buffer
pH 7.0 at room temperature for 3.5 h. The ratio of catechol
and aniline was 1 to 2, and 100 U of laccase and 924 U of
Entry
R1
Yield[a] (%) of product
Without lipase With lipase
1
2
3
4
5
H
H
OCH3
Cl
CH3
23
30
23
25
30
32
28[b]
37
51
50
[a] Isolated yield. [b] Used 1848U of lipase PS.
After the preliminary study, the reaction between cate-
lipase per 1 mmol of catechol were used. An insoluble red chol and other anilines was conducted. The results of these
color product precipitated out of solution during the reac- studies are summarized in Table 4. The reaction of catechol
tion. Therefore, the product was readily collected by fil- and anilines in the presence of laccase and lipase PS pro-
tration completion of reaction. The results show that the vided a higher yield than the reaction in presence of laccase
yield of the reaction with lipase PS increased by ca. 30% only. The yield of the product in the reaction with lipase PS
when compare to the yield of the reaction without lipase increased up to 70% compare to the reaction without lipase
PS. Next, the amount of lipase PS used in the reaction was PS (Table 4, Entry 4). Therefore, the overall yield of the
increased from 924 U to 1848 U per 1 mmol of catechol to product of this reaction system can be enhanced by lipase
study the effect of lipase dose on the reaction system. This PS.
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S. Witayakran, A. J. Ragauskas
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temperature for 4 h. After the reaction was completed, the reaction
mixture was extracted by EtOAc. The organic phase was combined,
dried with MgSO4, and the solvents evaporated. The resulting
crude products were purified by silica column chromatography,
using EtOAc and petroleum ether as an eluent to obtain 3-acetyl-
5,6-dihydroxy-2-methylbenzofuran (3a). Products were charac-
Conclusion
This study demonstrates the potential of using lipase to
catalyze the Michael addition reaction, and presents a new
cocatalytic enzymatic system employing laccase and lipase
for green chemistry synthesis. Lipase was found to catalyze
the addition reaction between laccase-generated o-quinones
and 1,3-dicarbonyl compounds in aqueous medium. In this
reaction, the catalytic system of laccase and lipase PS was
regioselective providing only one isomer product and is the
first example of a two enzyme catalytic system for the syn-
thesis of benzofurans. The yields of the products from reac-
tion were shown to depend on the reactivity of the starting
catechols and β-dicarbonyl compounds. Based on our ex-
perimental results, catechols with moderate reactivity yield
benzofuran products in excellent yield. Moreover, lipase
was also shown to catalyze the addition reaction between
laccase-generated o-quinone and aromatic amines. In the
presence of lipase and laccase, the yield of the final prod-
ucts increased in the range from 30 to 70% when compare
to the reaction in the presence of laccase alone. Therefore,
this paper illustrates a unique aqueous-based two-enzyme
system for green chemistry synthesis and future applications
are under study.
1
terized by H NMR, 13C NMR and MS.
Procedure for the Study of the Reaction of 1a and 2a (with and with-
out Lipase PS): In a 250-mL round-bottom flask, 40 mL of 0.10 
phosphate buffer pH 7.0 and 1a (2 mmol, 0.2202 g) were mixed
together. Next, 200 U of laccase was added to reaction mixture and
then, 2a (0.4004 g, 410 µL, 4 mmol) and 1848 U of lipase PS (or
no lipase) were added. The reaction was then stirred at room tem-
perature in a flask open to the atmosphere for 4.5 h. A 3 mL ali-
quot of the reaction mixture was taken every 30 min during the
reaction and extracted with 10 mL of EtOAc. The organic phase
was then dried with MgSO4, filtered and concentrated under re-
duced pressure. The resulting crude product was submitted to
1
quantitative H NMR analysis to measure the formation of prod-
uct 3a using [D6]DMSO as solvent and 1,3,5-trioxane as internal
standard.
General Procedure for the Reaction Between Catechols and Anilines:
In a 250-mL round-bottom flask, 30 mL of 0.10  phosphate
buffer pH 7.0 and 1a (0.1101 g, 1 mmol) were mixed together. Next,
100 U of laccase was added to reaction mixture and then, 4a
(0.1862 g, 182 µL, 2 mmol) and 924 U of lipase PS were added. The
reaction was then stirred under air at room temperature for 3.5 h.
After the reaction was finished, the reaction mixture was filtered
to collect the red color product 5a as red solid (87 mg, 30%). Prod-
Experimental Section
1
1
ucts were characterized by H NMR, 13C NMR and MS. H and
13C assignments and HMBC correlation for compound 5b, 5c, and
5d are summarized in Table 5.
General Information: All chemicals were used as received without
further purification. Laccase (EC 1.10.3.2) from Trametes Villosa
was donated by Novo Nordisk Biochem, North Carolina. Lipases
were purchased from Aldrich. Unit definition of each lipase is dif-
ferent depending on the method that Aldrich used to measure li-
Compound 5b: Red solid, yield 129.5 mg (37%), m.p. 161–162 °C.
1H NMR (400 MHz, CDCl3): δ = 8.50 (br. s, 1 H, OH), 7.56 (br.
s, 1 H, OH), 7.08 (d, J = 7 Hz, 4 H, 4ϫCH arom.), 6.94 (d, J =
8 Hz, 4 H, 4ϫCH arom.), 6.07 (s, 2 H, 2ϫCH), 3.84 (s, 6 H,
2ϫOCH3) ppm. 13C NMR (100 MHz, CDCl3): δ = 157.4, 151.9,
135.3, 123.9, 114.2, 95.1, 55.1 ppm. IR (KBr): ν¯ = 3293 (s), 3246
(s), 3040 (w), 2834 (w), 1739 (w), 1654 (w), 1606 (s), 1580 (s), 1525
(s), 1511 (s), 1411 (s), 1330 (m), 1286 (m), 1244 (s), 1217 (s), 1199
(s), 1173 (m), 1033 (m), 840 (m) cm–1. MS (EI): m/z = 350 (86)
[M+], 319 (100), 291 (12), 174 (15), 146 (12), 92 (7), 77 (9). HRMS
(EI): calcd. for C20H18N2O4 350.1266; found 350.1247.
pase activity. The enzymes were kept frozen until used. 1H and 13
C
NMR spectra were recorded on a Bruker-400 spectrometer op-
erating at 400 MHz for H and 100 MHz for 13C in [D6]DMSO or
1
CDCl3 using tetramethylsilane (TMS) as the internal standard. All
reactions were monitored by TLC. TLC was performed on alumi-
num sheets precoated with silica gel 60 F254 (EMD Chemicals).
Column chromatography was performed on Combiflash Compan-
ion instrument (Teledyne Isco company) using RediSep normal-
phase flash columns. Mass spectra were carried out in The Georgia
Institute of Technology Bioanalytical Mass Spectrometry Facility.
3a,[9] 3b,[10] 3c,[9] 3d,[6l] and 5a[11] are known compounds and our
1H and 13C NMR spectroscopic data are consistent with those in
the literature.
Compound 5c: Red solid, yield 182.6 mg (51%), m.p. 219–221 °C.
1H NMR (400 MHz, [D6]DMSO): δ = 9.24 (br. s, 2 H, 2ϫOH),
7.44 (d, J = 7 Hz, 4 H, 4ϫCH arom.), 7.19 (br. s, 4 H, 4ϫCH
arom.), 5.81 (s, 2 H, 2ϫCH) ppm. 13C NMR (100 MHz, [D6]-
DMSO): δ = 152.0, 142.6, 129.1, 128.9, 123.7, 97.5z ppm. IR
(KBr): ν¯ = 3298 (s), 3031 (w), 1736 (w), 1660 (w), 1606 (m), 1573
(s), 1536 (s), 1493 (s), 1480 (s), 1415 (s), 1334 (s), 1221 (s), 1188 (s),
1087 (m), 1007 (m), 830 (m) cm–1. MS (EI): m/z = 358 (M+, 42%),
323 (80), 288 (8), 178 (18), 144 (15), 127 (100), 84 (57), 65 (18),
49 (75). HRMS (EI): calcd. for C18H12N2O2Cl2 358.0275; found
358.0266.
Compound 5d: Red solid, yield 159 mg (50%), m.p. 194–196 °C. 1H
NMR (400 MHz, CDCl3): δ = 8.55 (br. s, 1 H, OH), 7.55 (br. s, 1
H, OH), 7.20 (d, J = 7 Hz, 4 H, 4ϫCH arom.), 7.02 (br. s, 4 H,
4ϫCH arom.), 6.09 (s, 2 H, 2ϫCH), 2.37 (s, 6 H, 2ϫCH3) ppm.
13C NMR (100 MHz, CDCl3): δ = 152.2, 135.5, 129.9, 122.0, 95.7,
20.9 ppm. IR (KBr): ν¯ = 3297 (s), 3031 (w), 2917 (w), 1739 (m),
1663 (w), 1600 (s), 1572 (s), 1533 (s), 1511 (s), 1488 (s), 1413 (s),
1337 (s), 1219 (s), 1189 (s), 1153 (s), 897 (m), 814 (m), 732 (m)
Enzyme Assay: Laccase used was from Trametes Villosa (EC
1.10.3.2). Laccase activity was determined by oxidation of 2,2Ј-
azinobis(3-ethylbenzylthiozoline-6-sulfonate) (ABTS).[12] The assay
mixture contained 25 µ ABTS, 0.10  sodium acetate (pH 5.0),
and a suitable amount of enzyme. The oxidation of ABTS was
followed by an absorbance increase at 420 nm (ε420
=
3.6ϫ104 –1 cm–1). Enzyme activity was expressed in units (U =
µol of ABTS oxidized per minute).
General Procedure for the Reaction Between Catechols and 1,3-Di-
carbonyl Compounds: In a 250-mL round-bottom flask, 30 mL of
0.10  phosphate buffer pH 7.0 and 1a (0.1101 g, 1 mmol) were
mixed together. Next, 100 U of laccase was added to reaction mix-
ture and then, 2a (0.2002 g, 205 µL, 2 mmol) and 924 U of lipase
PS were added. The reaction was then stirred under air at room
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Cocatalytic Enzyme System for Michael Addition Reaction
1
Table 5. H and 13C assignments and HMBC correlation for com-
[3] a) D. Gygax, P. Spies, T. Winkler, U. Pfaar, Tetrahedron 1991,
47, 5119–5122; b) J. Thiem, T. Wiemann, Angew. Chem. Int.
Ed. Engl. 1991, 30, 1163–1164; c) G. F. Herrmann, Y. Ichikawa,
C. Wandrey, F. C. A. Gaeta, J. C. Paulson, C.-H. Wong, Tetra-
hedron Lett. 1993, 34, 3091–3094; d) A. I. Scott, Synlett 1994,
871–883; e) V. Ken, J. Thiem, Angew. Chem. Int. Ed. Engl. 1995,
34, 893–895; f) P. J. Duggan, E. Parker, J. Coggins, C. Abell,
Bioorg. Med. Chem. Lett. 1995, 5, 2347–2352; g) R. Fernández-
Lafuente, J. Guisán, M. Pregnolato, M. Terreni, Tetrahedron
Lett. 1997, 38, 4693–4696; h) O. H. Justiz, R. Fernandez-Laf-
uente, J. M. Guisan, P. Negri, G. Pagani, M. Pregnolato, M.
Terreni, J. Org. Chem. 1997, 62, 9099–9106; i) H.-S. Bae, S.-G.
Lee, S.-P. Hong, M.-S. Kwak, N. Esaki, K. Soda, M.-H. Sung,
J. Mol. Catal. B 1999, 6, 241–247; j) M. A. Wegman, L. M.
van Langen, F. van Rantwijk, R. A. Sheldon, Biotechnol. Bio-
eng. 2002, 79, 356–361.
pound 5b, 5c, and 5d.[a]
Compound 5b
C atom[b]
13C (δ)
1H (δ)
1H-13C correlation
2, 2Ј
95.1
6.07, s, 2 H
C3, C3Ј
5, 5Ј, 9, 9Ј
123.9
7.08, d, 4 H (7)
C4, C4Ј, C6, C6Ј, C7,
C7Ј, C8, C8Ј
6, 6Ј, 8, 8Ј
114.2
55.1
6.94, d, 4 H (8)
C4, C4Ј, C5, C5Ј, C7,
C7Ј, C9, C9Ј
C7, C7Ј
[4] B. Seisser, I. Lavandera, K. Faber, J. H. L. Spelberg, W. Krou-
til, Adv. Synth. Catal. 2007, 349, 1399–1404.
10, 10Ј
3.84, s, 6 H
[5] a) S. G. Burton, Curr. Org. Chem. 2003, 7, 1317–1331; b) H.
Claus, Micron 2004, 35, 93–96; c) S. Nicotra, A. Intra, G. Otto-
lina, S. Riva, B. Danieli, Tetrahedron: Asymmetry 2004, 15,
2927–2931; d) S. Nicotra, M. R. Cramarossa, A. Mucci, U. M.
Pagnoni, S. Riva, L. Forti, Tetrahedron 2004, 60, 595–600; e)
X. Zhang, G. Eigendorf, D. W. Stebbing, S. D. Mansfield, J. N.
Saddler, J. N, Arch. Biochem. Biophys. 2002, 405, 44–54; f) A.
Leonowicz, N. S. Cho, J. Luterek, A. Wilkolazka, M. Wojtas-
Wasilewska, A. Matuszewska, M. Hofrichter, D. Wesenberg, J.
Rogalski, J. Basic Microbiol. 2001, 41, 185–227; g) C. Galli, P.
Gentili, J. Phys. Org. Chem. 2004, 17, 973–977; h) F. d’Acunzo,
C. Galli, P. Gentili, F. Sergi, New J. Chem. 2006, 30, 583–591; i)
O. V. Morozova, G. P. Shumakovich, M. A. Gorbacheva, S. V.
Shleev, A. I. Yaropolov, Biochemistry (Moscow) 2007, 72,
1136–1150.
[6] a) T. H. Niedermeyer, A. Mikolasch, M. Lalk, J. Org. Chem.
2005, 70, 2002–2008; b) S. Riva, Trends Biotechnol. 2006, 24,
219–226; c) S. Ciecholewski, E. Hammer, K. Manda, G. Bose,
V. T. H. Nguyen, P. Langer, F. Schauer, Tetrahedron 2005, 61,
4615–4619; d) M. Kurisawa, J. E. Chung, H. Uyama, S. Kobay-
ashi, Biomacromolecules 2003, 4, 1394–1399; e) A. Mikolasch,
E. Hammer, U. Jonas, K. Popowski, A. Stielow, F. Schauer,
Tetrahedron 2002, 58, 7589–7593; f) M. Hosny, J. Rosazza, J.
Agric. Food Chem. 2002, 50, 5539–5545; g) A. Schäfer, M.
Specht, A. Hetzheim, W. Francke, F. Schauer, Tetrahedron
2001, 57, 7693–7699; h) K. Manda, E. Hammer, A. Mikolasch,
T. Niedermeyer, J. Dec, A. D. Jones, A. J. Benesi, F. Schauer,
J.-M. Bollag, J. Mol. Catal. B: Enzym. 2005, 35, 86–92; i) H.
Leutbecher, J. Conrad, I. Klaiber, U. Beifuss, Synlett 2005,
3126–3130; j) S. Witayakran, A. Zettili, A. J. Ragauskas, Tetra-
hedron Lett. 2007, 48, 2983–2987; k) S. Witayakran, A. J. Ra-
gauskas, Green Chem. 2007, 9, 475–480; l) S. Witayakran, L.
Gelbaum, A. J. Ragauskas, Tetrahedron 2007, 63, 10958–10962.
[7] a) S. H. Krishna, N. G. Karanth, Catal. Rev. 2002, 44, 499–
591; b) R. Sharma, Y. Chisti, U. C. Banerjee, Biotechnol. Adv.
2001, 19, 627–662; c) F. Hasan, A. A. Shah, A. Hameed, En-
zyme Microb. Technol. 2006, 39, 235–251.
OH (1, 1Ј)
7.56, br. s, 1 H
8.50, br. s, 1 H
Compound 5c
C atom[c]
13C (δ)
1H (δ)
1H-13C correlation
2, 2Ј
97.5
129.1
123.7
5.81, s, 2 H
C3, C3Ј
C4, C4Ј, C6, C6Ј, C8, C8Ј
C5, C5Ј, C7, C7Ј, C9, C9Ј
5, 5Ј, 9, 9Ј
6, 6Ј, 8, 8Ј
OH (1, 1Ј)
7.44, d, 4 H (7)
7.19, br. s, 4 H
9.24, br. s, 2 H
Compound 5d
C atom[d]
13C (δ)
1H (δ)
1H-13C correlation
2, 2Ј
5, 5Ј, 9, 9Ј
6, 6Ј, 8, 8Ј
95.7
122.0
129.9
6.09, s, 2 H
7.02, br. s, 4 H
7.20, d, 4 H (7)
C3, C3Ј
C6, C6Ј, C7, C7Ј, C8, C8Ј
C5, C5Ј, C9, C9Ј, C10,
C10Ј
10, 10Ј
OH (1, 1Ј)
20.9
2.37, s, 6 H
7.55, br. s, 1 H
8.55, br. s, 1 H
C6, C6Ј, C7, C7Ј, C8, C8Ј
[a] Measure in CDCl3 or [D6]DMSO at 100 MHz (13C) or 400 MHz
[1H, J (Hz) values in parentheses]. Chemical shifts are express in δ
(ppm). [b] Compound 5b: 13C (δ) of C-3/3Ј, C-4/4Ј and C7/7Ј =
151.9, 135.3, and 157.4 ppm. [c] Compound 5c: 13C (δ) of C-3/3Ј,
C-4/4Ј and C7/7Ј = 152.0, 142.6, and 128.9 ppm. [d] Compound
5d: 13C (δ) of C-3/3Ј and C7/7Ј = 152.2 and 135.5 ppm.
cm–1. MS (EI): m/z = 318 (M+, 42%), 303 (100), 275 (15), 158 (13),
130 (8), 91 (14), 65 (8), 49 (11). HRMS (EI): calcd. for C20H18N2O2
318.1368; found 318.1348.
Acknowledgments
[8] a) O. Torre, I. Alfonso, V. Gotor, Chem. Commun. 2004, 1724–
1725; b) M. Svedendahl, K. Hult, P. Berglund, J. Am. Chem.
Soc. 2005, 127, 17988–17989; c) H.-X. Dai, S.-P. Yao, J. Wang,
Biotechnol. Lett. 2006, 28, 1503–1507.
The authors acknowledge financial support from the IPST@GT
student fellowship and the Royal Thai Government Scholarship.
[9] D. Nematollahi, M. Rafiee, J. Electroanal. Chem. 2004, 566,
[1] P. Anastas, J. Warner in Green Chemistry: Theory and Practice,
Oxford University Press, New York, 1998.
31–37.
[10] D. Nematollahi, M. Rafiee, Green Chem. 2005, 7, 638–644.
[11] S. Uchiyama, Y. Hasebe, J. Nishimoto, H. Hamana, Y. Maeda,
Y. Yoshida, Electroanalysis 1998, 10, 647–650.
[12] R. Bourbonnais, D. Leech, M. G. Paice, Biochim. Biophys. Acta
1998, 1379, 381–390.
[2] a) S. F. Mayer, W. Kroutil, K. Faber, Chem. Soc. Rev. 2001, 30,
332–339; b) S. M. Glueck, S. F. Mayer, W. Kroutil, K. Faber,
Pure Appl. Chem. 2002, 74, 2253–2257; c) H. Pellissier, Tetrahe-
dron 2006, 62, 2143–2173; d) L. F. Tietze, G. Brache, K. M.
Gericke in Domino Reactions in Organic Synthesis, Wiley-VCH,
Weinheim, 2006, pp. 529–541.
Received: August 13, 2008
Published Online: Novembver 28, 2008
Eur. J. Org. Chem. 2009, 358–363
© 2009 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.eurjoc.org
363