Ethyl acetate as an acyl donor in the continuous flow kinetic resolution of (±)-1-phenylethylamine catalyzed by lipases

Article abstract of DOI:10.1039/c3ob40437d

The synthesis of chiral amines is still a challenge for organic synthesis since optically pure amines are of great importance for the pharmaceutical and agrochemical industries. Among all the methodologies developed until now, chemoenzymatic dynamic kinetic resolution has proven to be useful for the preparation of enantioenriched primary chiral amines. In our continuous efforts toward the development of a continuous flow process, herein we report our results on the continuous flow kinetic resolution of (±)-1- phenylethylamine leading to the desired products with high enantiomeric ratios (>200) and short residence times (40 minutes) using ethyl acetate as the acyl donor.

Full text of DOI:10.1039/c3ob40437d

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Ethyl acetate as an acyl donor in the continuous flow
kinetic resolution of ( )-1-phenylethylamine catalyzed
by lipases†
Cite this: DOI: 10.1039/c3ob40437d
Amanda S. de Miranda, Leandro S. M. Miranda and Rodrigo O. M. A. de Souza*
The synthesis of chiral amines is still a challenge for organic synthesis since optically pure amines are of
great importance for the pharmaceutical and agrochemical industries. Among all the methodologies
developed until now, chemoenzymatic dynamic kinetic resolution has proven to be useful for the prepa-
ration of enantioenriched primary chiral amines. In our continuous efforts toward the development of
a continuous flow process, herein we report our results on the continuous flow kinetic resolution of
( )-1-phenylethylamine leading to the desired products with high enantiomeric ratios (>200) and short
residence times (40 minutes) using ethyl acetate as the acyl donor.
Received 18th December 2012,
Accepted 11th March 2013
DOI: 10.1039/c3ob40437d
www.rsc.org/obc
1. Introduction
Optically pure amines, in which the nitrogen atom is bound to
a chiral center (chiral amine), are of great importance for the
pharmaceutical and agrochemical industries, being used as
resolving agents, chiral auxiliaries, ligands and intermediates
in the synthesis of many drugs and pesticides.1
The synthesis of chiral amines is still an unsolved synthetic
challenge. Over the years, several methods have been develo-
ped, including the multi-step protocol of carbanions addition
to chiral sulphinyl, sulphonyl or phosphoryl imines,2–4 the
hydrogenation of acyl enamines with chiral phosphorous
ligands,5–8 transition metal or organocatalyzed or enzymatic
(transaminases) asymmetric imine reduction,9–19 asymmetric
hydroamination of olefins,16,20–22 crystallization,23–25 and enzy-
matic catalyzed kinetic or dynamic resolution.23,26–33 All of
these approaches suffer from limited scope and greenness,
and no definitive approach exists.
Among them, chemoenzymatic dynamic kinetic resolution
has proven to be useful for the preparation of enantioenriched
primary chiral amines due to its low operational and catalyst
cost and in general more suitable process regarding green
chemistry principles. In the kinetic resolution mediated by
lipases, a racemate (1) is resolved through enantioselective
enzymatic acylation, providing enantioenriched amide (3) and
amine (2) with opposite configuration.27,30,34–38 If only one
enantiomer is desired, a metal catalyzed racemization reaction
Scheme 1 Kinetic dynamic resolution of racemic amines.
can be coupled with the kinetic resolution in a one-pot pro-
cedure, named dynamic kinetic resolution (Scheme 1).39–41
Because of their stability, broad substrate range, non-
requirement of co-factors, compatibility with organic solvents
and mild reaction conditions, lipases are the most used
enzymes for kinetic resolution. In addition, many lipases are
readily available and can be immobilized and reutilized, being
suitable for industrial purposes. Among the lipases, Candida
antarctica lipase B (CALB) is by far the most used enzyme in
the kinetic resolution of amines. As acyl donors, many esters
have been used, alkyl 2-methoxyacetates esters being very
common.32,35,42,43
Kinetic resolution of amines with immobilized lipases has
been reported to occur at temperatures between 25 °C and
70 °C and reaction times varying from 4 h to 72 h, depending
on the nature of the acyl donor (i.e. vinyl acetate, isopropenyl
acetate and 2-methoxy ethyl acetate). Just a few examples report
the use of ethyl acetate as an acyl donor and in most cases long
reaction times and moderate yields are obtained, mainly due to
the low reactivity of this ester in the acylation reaction.34,44
Continuous flow processes present many advantages over
batch procedures, which include: precise control over reaction
Biocatalysis and Organic Synthesis Group, Federal University of Rio de Janeiro,
Chemistry Institute, Rio de Janeiro, CEP22941-909, Brazil.
E-mail: rodrigosouza@iq.ufrj.br
†Electronic supplementary information (ESI)
available. See DOI:
10.1039/c3ob40437d
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conditions, such as time, temperature and pressure, leading to temperatures (25–70 °C) and residence times using a column
more reproducible processes; enhanced mass transfer; safety; reactor packed with Novozyme 435. Concerning the residence
ease of product isolation; possibility of automation and ease of times, it was observed that short residence times lead to a
scale-up.45–47
decrease in conversion since the time of contact between the
To the best of our knowledge only a few examples of kinetic substrate and the enzyme is reduced (see ESI† for further
resolution of amines under continuous flow conditions have details). The best residence time observed during the screen-
been reported in the literature using lipases as catalysts.48,49 ing process was that of 40 minutes (Table 1).
Roengpithya and co-workers reported a continuous process for
The influence of the acylating agent on reaction conversion
dynamic kinetic resolution of ( )-1-phenylethylamine using a and enantiomeric ratio is depicted in Table 1, under different
membrane assisted two vessels system, but the reaction took temperatures. As can be seen, isopropenyl acetate seems to be
72 h and gave poor isolated yields.49 BASF has also used the the most reactive acylating agent, leading to low enantiomeric
continuous flow approach for the kinetic resolution of 1-phenyl- ratios (E) at all temperatures studied (entries 1–4, Table 1).
ethylamine using Burkholderia plantarii lipase immobilized on Besides that, formation of an imine derivative probably
polyacrylate and 2-methoxy ethyl acetate as the acylating agent. obtained from the reaction between the free amine and propa-
High ee values were obtained but a long residence time is none (derived from the isopropenyl ester) was also detected by
needed (5–7 h) to achieve satisfactory results.48
GC-MS, and also contributed to the low yields obtained.
Alkyl 2-methoxyacetates have been also used as acyl donors
In our continuous efforts toward developing continuous
flow biocatalyzed reactions50–57 herein we report our results on in the kinetic resolution of amines. These esters are less reac-
the kinetic resolution of ( )-1-phenylethylamine (1) catalyzed tive than isopropenyl acetate but much more reactive than
by Candida antarctica lipase B immobilized on acrylic resin simple esters, such as ethyl acetate, and they are often used
(Novozyme 435) using a continuous flow reactor and ethyl because of the milder conditions required for kinetic resolu-
acetate as the acyl donor.
tion and easier hydrolysis of the amide product.33,37,55,57 The
results obtained by the use of methyl 2-methoxyacetate as the
acyl donor in the kinetic resolution of ( )-1-phenylethylamine
(1) show that the desired product can be obtained with high
conversions and enantiomeric ratios as expected, even at room
2. Results and discussion
We began our study screening the acylating agents (isoprope- temperature (entry 5, Table 1). The increase in reaction temp-
nyl acetate, methyl 2-methoxyacetate, ethyl acetate) at different erature resulted in a lower enantiomeric ratio, as observed in
Table 1 Kinetic resolution of ( )-1-phenylethylamine (1) under continuous flow conditions
Entry
Temp. (°C)
Acyl donor
Conv.a (%)
Eb
1
2
3
4
5
6
7
8
25
35
50
70
25
35
50
70
25
35
50
70
25
35
50
70
Isopropenyl acetate
Isopropenyl acetate
Isopropenyl acetate
Isopropenyl acetate
2-Methoxy methylacetate
2-Methoxy methylacetate
2-Methoxy methylacetate
2-Methoxy methylacetate
Ethyl acetate
54
50
51
57
49
50
51
53
31
38
44
48
35
37
43
50
28
24
11
9
>200
>200
146
83
>200
>200
>200
>200
168
87
9
10
11
12
13
14
15
16
Ethyl acetate
Ethyl acetate
Ethyl acetate
Ethyl acetatec
Ethyl acetatec
Ethyl acetatec
84
58
Ethyl acetatec
Reaction conditions: To an Omnifit column packed with 1.4 g of Novozyme 435 (volume = 3.927 mL), a solution (0.080 M) of ( ) of
1-phenylethylamine and 0.320 M of acyl donor in toluene were pumped through the column at a flow rate of 0.1 mL min−1 (residence time =
40 min).a Conversions (C) were calculated as C = eer/(eep + eer), where eer and eep are the enantiomeric excess of amine 2 and amide 3,
respectively. b Enantiomeric ratios (E) were calculated as E = ln[1 − C(1 − eep)/ln[1 − C(1 + eep)]. Enantiomeric excess was determined by GC.
c Ethyl acetate was used as the solvent. All experiments were performed with the same enzyme activity.
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entry 8 (Table 1). Since methyl 2-methoxyacetate is a relatively
expensive acyl donor, we also carried out the kinetic resolution
of ( )-1-phenylethylamine (1) by varying the acyl donor :
substrate ratio and we found that at room temperature it has
little influence on enantiomeric ratio and conversion, leading
to excellent enantiomeric ratios (>200) with an acyl donor :
substrate ratio of 1 : 1 and a residence time of 40 minutes.
Ethyl acetate was studied as the acyl donor in order to
reduce process cost (ethyl acetate is up to 20-fold cheaper than
2-methoxy methylacetate51) and work with a more readily avail-
able starting material. Surprisingly, ethyl acetate presented
interesting behavior under continuous flow conditions allow-
ing us to achieve excellent results under mild temperature
(50 °C) (entries 11 and 12, Table 1). The reaction under con-
tinuous flow was found to occur fast and with high conver-
sions and enantiomeric ratios. The space-time yield obtained
for this transformation, 0.25 kg m3 h−1, indicates that
this process can be an alternative to the traditional batch
reaction.52,58,59
In order to access a greener process we also performed the
kinetic resolution of ( )-1-phenylethylamine (1) in ethyl acetate
as the solvent. High conversions were obtained at 50 °C and
70 °C with residence times of 40 minutes, but enantiomeric
ratios were much lower in comparison to kinetic resolution
performed in toluene (entries 15 and 16, Table 1), indicating a
concurrent uncatalyzed acylation with such a high concen-
tration of the acyl donor.
Fig. 1 Recyclability of the continuous flow system.
Table 2 Kinetic resolution of ( )-1-phenylethylamine (1) using ethyl acetate as
the acyl donor
Entry
Conc. (mM)
Conv.a (%)
Eb
1
2
3
4
5
6
7
80
150
300
450
600
750
900
48
46
43
41
44
42
42
>200
>200
>200
>200
>200
>200
>200
Reaction conditions: To an Omnifit column packed with 1.4 g of
Novozyme 435 (volume = 3.927 mL), a solution (X M) of ( ) of
1-phenylethylamine and 0.320 M of acyl donor in toluene were
pumped through the column at a flow rate of 0.1 mL min−1 (residence
time = 40 min).a Conversions (C) were calculated as C = eer/(eep + eer),
where eer and eep are the enantiomeric excess of amine 2 and amide 3,
respectively. b Enantiomeric ratios (E) were calculated as E = ln[1 −
C(1 − eep)/ln[1 − C(1 + eep)]. Enantiomeric excess was determined by GC.
In addition, we have carried out the kinetic resolution of
( )-1-phenylethylamine (1) and ethyl acetate with other com-
mercially available immobilized enzymes from Rhizomucor
miehei (Lipozyme RM IM), Thermomyces lanuginosus (Lipozyme
TL IM) and Burkholderia cepacia (PS Amano IM), but none was
found to provide the product with conversions higher than
10% at 50° or 70 °C with residence times of 40 minutes (see
ESI† for further details).
The recyclability of the immobilized CALB was also evalu-
ated as well as the stability of the continuous flow system
during the kinetic resolution of ( )-1-phenylethylamine (1)
using ethyl acetate as the acetylating agent at 70 °C and
40 min of residence time. The packed bed system was stable
for more then 9 hours of operation. The stability is shown in
Fig. 1 where samples were taken every hour and analyzed to
verify the conversion and enantiomeric excess of the product.
Next we assayed increasing concentrations of stock solu-
tions containing starting materials from 0.080 M to 0.9 M in
order to verify the capabilities of the continuous flow system to
operate with highly concentrated media and some selected
results are presented in Table 2. The results show that
( )-1-phenylethylamine (1) can be converted to optically pure
amide (3) and amine (2) under continuous flow at concen-
obtained, increasing the attractiveness for large scale reactions
of the approach here developed.
In summary, we have presented an efficient and low cost
procedure for the kinetic resolution of ( )-phenylethylamine
under continuous flow conditions. In comparison to existing
batch processes, the kinetic resolution herein described pre-
sents shorter reaction time, higher productivity, easier work-
up and is more suitable to be automatized and scaled up for
industrial purposes. Currently, our group is working toward
the integration of this process with dynamic resolution under
continuous flow conditions.
Acknowledgements
trations as high as 0.9 M (with no increase in the bed volume). We thank CAPES (Coordenação de Aperfeiçoamento de Pessoal
To the best of our knowledge no precedent exists in the litera- de Nível Superior), CNPq (Conselho Nacional de Desenvolvi-
ture where a kinetic resolution of amines is run under such mento Científico e Tecnológico), FAPERJ (Fundação Carlos
concentration. The use of concentrated solutions increases the Chagas Filho de Amparo a Pesquisa do Estado do Rio de
productivity from 0.055 to 0.620 g h−1 g of immobilized Janeiro) and FINEP (Agência Financiadora de Estudos e Proje-
enzyme−1, an 11-fold increase for each one of the products tos) for financial support.
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