Lipase-catalyzed enantioselective ring opening of unactivated alicyclic-fused beta-lactams in an organic solvent.

Article abstract of DOI:10.1021/ol034096o

[reaction: see text] A highly efficient and very simple method was developed for the synthesis of enantiopure beta-amino acids (e.g. cispentacin) and beta-lactams through the enzyme-catalyzed enantioselective ring opening of unactivated alicyclic beta-lac

Full text of DOI:10.1021/ol034096o

ORGANIC
LETTERS
2003
Vol. 5, No. 8
1209-1212
Lipase-Catalyzed Enantioselective Ring
Opening of Unactivated Alicyclic-Fused
â-Lactams in an Organic Solvent
Eniko Forro´ and Ferenc Fu1lo1p*
Institute of Pharmaceutical Chemistry, UniVersity of Szeged, P.O. Box 121,
H-6701 Szeged, Hungary
fulop@pharma.szote.u-szeged.hu
Received January 20, 2003
ABSTRACT
A highly efficient and very simple method was developed for the synthesis of enantiopure â-amino acids (e.g. cispentacin) and â-lactams
through the enzyme-catalyzed enantioselective ring opening of unactivated alicyclic â-lactams in organic media. High enantioselectivity (E >
200) was observed when the Lipolase (lipase B from Candida antarctica)-catalyzed reactions were performed with H O (1 equiv) in diisopropyl
2
ether at 60 °C. The resolved products, obtained in good chemical yield (36−47%), could be easily separated.
The â-amino acids and â-lactams are of biological and
chemical importance. Some â-amino acids themselves exhibit
antibacterial activity (e.g. cispentacin).1 They can serve as
precursors of â-lactams,2 and can be used as building blocks
for the synthesis of modified peptides with increased activity
and stability,3 and in drug research.4 Well-defined three-
dimensional structures (e.g. â-peptides with possible anti-
biotic activity) similar to those of natural peptides can also
be formed from â-amino acids.5 Alicyclic â-amino acids can
be used in heterocyclic6 and combinatorial7 chemistry.
Besides the utility of monocyclic â-lactams as antibiotic
agents (e.g. monobactams), they are used as intermediates
in â-amino acid synthesis,8 and as building blocks for short
peptide segments,9 taxoid antitumor agents,10 alkaloids,11
heterocycles12 and other types of compounds of biological
and medicinal interest. Consequently, in the past few years
(5) (a) Seebach, D.; Matthews, J. L. Chem. Commun. 1997, 2015-2022.
(b) Appela, D. H.; Christianson, L. A.; Klein, D. A.; Powell, D. R.; Huang,
X. L.; Barchi, J. J.; Gellman, S. H. Nature 1997, 387, 381-384. (c) Gellman,
S. H. Acc. Chem. Res. 1998, 31, 173-180. (d) Porter, E. A.; Wang, X. F.;
Lee, H. S.; Weisblum, B.; Gellman, S. H. Nature 2000, 404, 565-565. (e)
Gu¨nther, R.; Hofmann, H.; Kuczera, K. J. Phys. Chem. B 2001, 105, 5559-
5567. (f) Martinek, T.; To´th, G. K.; Vass, E.; Hollo´si, M.; Fu¨lo¨p, F. Angew.
Chem., Int. Ed. 2002, 41, 1718-1721.
(6) (a) Szakonyi, Z.; Martinek, T.; Hete´nyi, A.; Fu¨lo¨p, F. Tetrahedron:
Asymmetry 2000, 11, 4571-4579. (b) Davies, S. G.; Bhalay G. Tetra-
hedron: Asymmetry 1996, 7, 1595-1596.
(7) Gedey, S.; Van der Eycken, J.; Fu¨lo¨p, F. Org. Lett. 2002, 4, 1967-
1969.
(8) (a) Ojima, I.; Delaloge, F. Chem. Soc. ReV. 1997, 26, 377-386. (b)
Palomo, C.; Aizpurua, J. M.; Ganboa, I.; Oiarbide, M. Synlett 2001, 1813-
1826.
(9) (a) Palomo, C.; Oiarbide, M.; Landa, A.; Esnal, A.; Linden, A. J.
Org. Chem. 2001, 66, 4180-4186. (b) Palomo, C.; Ganboa, I.; Oiarbide,
M.; Sciano, G. T.; Miranda, J. L. ArkiVoc 2002, 5, 8-16.
(10) Juaristi, E. EnantioselectiVe Synthesis of â-Amino Acids; Wiley-
VHC: New York, 1997.
(1) (a) Fu¨lo¨p, F. Chem. ReV. 2001, 101, 2181-2204. (b) Fu¨lo¨p, F. In
Studies in Natural Product Chemistry; Atta-ur-Rahman, Ed.; Elsevier
Science Publishers: New York, 2000; Vol. 22, pp 273-306. (c) Park, K.;
Kurth, M. J. Tetrahedron 2002, 58, 8629-8659.
(2) (a) Escalante, J.; Gonza´lez-Tototzin, M. A.; Avin˜a, J.; Mun˜oz-Mun˜iz,
O.; Juaristi, E. Tetrahedron 2001, 57, 1883-1890. (b) Krishnaswamy, D.;
Govande, V. V.; Gumaste, V. K.; Bhawal, B. M.; Deshmukh, A. R. A. S.
Tetrahedron 2002, 58, 2215-2225. (c) Sleeman, M. C.; MacKinnon, C.
H.; Hewitson, K. S.; Schofield, C. J. Bioorg. Med. Chem. Lett. 2002, 12,
597-599.
(3) (a) Brashear, K. M.; Hunt, C. A.; Kucer, B. T.; Duggan, M. E.;
Hartman, G. D.; Rodan, G. A.; Rodan, S. B.; Leu, C.; Prueksaritanont, T.;
Fernandez-Metzler, C.; Barrish, A.; Homnick, C. F.; Hutchinson, J. H.;
Coleman, P. J. Bioorg. Med. Chem. Lett. 2002, 12, 3483-3486. (b) Steer,
D. L.; Lew, R. A.; Perlmutter, P.; Smith, A. I.; Aguilar, M. I. Curr. Med.
Chem. 2002, 9, 811-822.
(4) (a) Fu¨lo¨p, F. Farmaco 2000, 55, 181-183. (b) Mittendorf, J.;
Kunisch, F.; Matzke, M.; Militzer, H. C.; Schmidth, A.; Scho¨nfeld, W.
Bioorg. Med. Chem. Lett. 2003, 13, 433-436. (c) Mittendorf, J.; Benet-
Buchholz, J.; Fey, P.; Mohrs, K. H. Synthesis 2003, 136-140.
(11) Wasserman, H. H.; Matsuyama, H.; Robinson, R. P. Tetrahedron
2002, 58, 7177-7190.
10.1021/ol034096o CCC: $25.00 © 2003 American Chemical Society
Published on Web 03/19/2003

a large number of syntheses (enantioselective synthesis and
enzymatic resolution) have been developed for enantiopure
alicyclic â-amino acid13 and â-lactam14-16 derivatives. One
good possibility for the preparation of enantiopure â-lactams
is an enantioselective enzyme-catalyzed hydrolysis of acti-
vated â-lactams.14 As an example, Adam et al. described the
simultaneous preparation of enantiopure amino acids and
â-lactams from R-methylene â-lactams by lipase-catalyzed
kinetic resolution through hydrolysis.14a Achilles et al.
described the enzyme-catalyzed hydrolysis of ethyl (()-2-
(2-oxo-4-phenylazetidin-1-yl)acetate with R-chymotrypsin.14b
However, these ring-opening methods have been limited to
activated â-lactams (activated with N-acyl-protecting groups
such as tert-butoxycarbonyl or acetyl). Evans et al. inves-
tigated the enzymatic ring-opening reactions of unactivated
(()-6-azabicyclo[3.2.0]-hept-3-en-7-one, and found that lac-
tamases in a special whole-cell preparation, ENZA-1 (Rhodo-
coccus equi NCIMB 40213), catalyzed the enantioselective
ring opening of this compound in water.15 Interestingly, the
saturated â-lactam remains intact under the same conditions.
An important indirect enzymatic route for enantiopure
alicyclic â-amino acid derivatives and â-lactams is through
lipase-catalyzed asymmetric acylation of the N-hydroxy-
methylated â-lactams or lipase-catalyzed hydrolysis of the
corresponding ester derivatives in an organic solvent.16,17
In this paper, we report a new direct method for the
enantioselective ring opening of unactivated cyclic â-lactams
(()-1-(()-4, yielding the ring-opened â-amino acids 1a-
4a and unreacted â-lactam enantiomers 1b-4b, which could
be easily separated (Scheme 1).
70 °C.14a Since they mentioned that the enzyme rapidly lost
activity at higher temperatures, we started our experiments
with Chirazyme L-2 (a carrier-fixed lipase B from Candida
antarctica) at 60 °C, but instead of water the ring-opening
reactions of (()-1-(()-4 were performed in diisopropyl
ether, using water (1 equiv) as nucleophile (Table 1, entries
Table 1. Conversion and Enantioselectivity of Ring Opening
of (()-1-(()-4
b
c
enzyme
H2O
conv ees eep
entry compda
(50 mg mL-1
)
(equiv) (%) (%) (%)
E
1d
2d
3d
4d
5d
6e
7e
8d
9e
10e
11e
12e
13e
14d
15d
16d
17d
(()-1
(()-1
(()-2
(()-2
(()-3
(()-3
(()-3
(()-3
(()-3
(()-3
(()-3
(()-3
(()-3
(()-3
(()-3
(()-4
(()-4
Chirazyme L-2
Lipolase
1
1
14
19
25
29
41
32
30
48
49
46
43
30
13
40
18
19
20
16 >95 >45
22 >95 >48
31 >95 >52
39 >95 >57
67 >95 >78
45 >95 >60
40 >95 >57
89 >95 >117
90 >95 >120
82 >95 >99
73 >95 >85
40 >95 >57
14 >95 >44
63 >95 >74
21 >95 >47
22 >95 >48
24 >95 >49
Chirazyme L-2
Lipolase
1
1
Chirazyme L-2
Chirazyme L-2
Chirazyme L-2
Lipolase
1
2
3
1
Lipolase
-
Lipolase
2
Lipolase
3
Lipolase
4
Lipolase
10
1
Novozyme 435
CAL-A
1
Chirazyme L-2
Lipolase
1
1
a 0.05 M substrate in diisopropyl ether, 60 °C. b According to GC.
c Calculated by using an internal standard (hexadecane). d After 20 h. e After
23 h.
1, 3, 5, and 16). High enantioselectivities but rather long
reaction times were observed. An additional enzyme screen-
ing including lipase PS (Pseudomonas cepacia), lipase AK
(Pseudomonas fluorescens), lipase AY (Candida rugosa),
Scheme 1. Enantioselective Ring Opening of Unactivated
Cyclic â-Lactams (()-1-(()-4
(13) (a) Faulconbridge, S. J.; Holt, K. E.; Sevillano, L. G.; Lock, C. J.;
Tiffin, P. D.; Tremayne, N.; Winter, S. Tetrahedron Lett. 2000, 41, 2679-
2681. (b) Bolm, C.; Dinter, C. L.; Sciffers, I.; Defre`re, L. Synlett 2001, 12,
1875-1877. (c) Yokota, Y.; Cortez, G. S.; Romo, D. Tetrahedron 2002,
58, 7075-7080. (d) Liu, M.; Sibi, M. P. Tetrahedron 2002, 58, 7991-
8035. (e) Aggarwal, V. K.; Roseblade, S. J.; Barrell, J. K.; Alexander, R.
Org. Lett. 2002, 4, 1227-1229. (f) Woll, M. G.; Fisk, J. D.; LePlae, P. R.;
Gellman, S. H. J. Am. Chem. Soc. 2002, 124, 12447-12452. (g) Porter, E.
A.; Wang, X. F.; Schmitt, M. A.; Gellman, S. H. Org. Lett. 2002, 4, 3317-
3319.
(14) (a) Adam, W.; Groer, P.; Humpf, H.-U.; Saha-Mo¨ller, C. R. J. Org.
Chem. 2000, 65, 4919-4922. (b) Achilles, K.; Schirmeister, T.; Otto, H.-
H. Arch. Pharm. Pharm. Med. Chem. 2000, 333, 243-253.
(15) Evans, C.; McCague, R.; Roberts, S. M.; Sutherland, A. G.; Wisdom,
R. J. Chem. Soc., Perkin Trans. 1 1991, 2276-2277.
The alicyclic â-lactams (()-1-(()-3 were prepared by
1,2-dipolar cycloaddition of chlorosulfonyl isocyanate (CSI)
to the corresponding cycloalkene,18 while (()-4 was obtained
by means of CSI cycloaddition to cyclooctadiene19 and
subsequent catalytic reduction in the presence of cyclohexene
as a hydrogen donor.17d
Adam et al. reported on the highly enantioselective
hydrolysis of the N-substituted â-lactam derivatives (E
usually >200) with the lipase Chirazyme L-2, in water at
(16) Nagai, H.; Shiozawa, T.; Achiwa, K.; Terao, Y. Chem. Pharm. Bull.
1993, 41, 1933-1938.
(17) (a) Csomo´s, P.; Kanerva, L. T.; Berna´th, G.; Fu¨lo¨p, F. Tetrahedron:
Asymmetry 1996, 7, 1789-1796. (b) Ka´ma´n, J.; Forro´, E.; Fu¨lo¨p, F.
Tetrahedron: Asymmetry 2000, 11, 1593-1600. (c) Fu¨lo¨p, F.; Palko´, M.;
Ka´ma´n, J.; La´za´r, L.; Sillanpa¨a¨, R. Tetrahedron: Asymmetry 2000, 11,
4179-4187. (d) Forro´, E.; AÄ rva, J.; Fu¨lo¨p, F. Tetrahedron: Asymmetry 2001,
12, 643-649.
(18) (a) Nativ, E.; Rona, P. Isr. J. Chem. 1972, 10, 55-58. (b) Singh,
R.; Cooper, R. D. G. Tetrahedron 1994, 50, 12049-12064. (c) Furet, P.;
Garcia-Echeverria, C.; Gay, B.; Scoepfer, J.; Zeller, M.; Rahuel, J. J. Med.
Chem. 1999, 42, 2358-2363.
(19) (a) Parsons, P. J.; Camp, N. P.; Underwood, J. M.; Harvey, D. M.
Tetrahedron 1996, 52, 11637-11642. (b) Parsons, P. J.; Camp, N. P.;
Edwards, N.; Sumoreeah, L. R. Tetrahedron 2000, 56, 309-315.
(12) (a) Cabell, L. A.; McMurray, J. S. Tetrahedron Lett. 2002, 43,
2491-2493. (b) Alcaide, B.; Almendros, P.; Alonso, J. M.; Aly, M. F.;
Pardo, C.; Saez, E.; Torres, M. R. J. Org. Chem. 2002, 67, 7004-7013.
1210
Org. Lett., Vol. 5, No. 8, 2003

Table 2. Lipolase-Catalyzed Ring Opening of (()-1-(()-4a
â-lactam (1b-4b)
â-amino acid (1a -4a )
time (h)
conv (%)
E
yieldb (%)
isomer
eec (%)
[R]25
yieldb (%)
isomer
eed (%)
[R]25
D
D
(()-1
(()-2
(()-3
(()-4
249
141
31
48
50
50
50
>200
>200
>200
>200
42
45
41
36
1S,5R
1S,6R
1S,7R
1S,8R
93
98
99
99
+35.9e
-3.6e
-5.1e
-18e
44
45
47
43
1R,2S
1R,2S
1R,2S
1R,2S
99
98
98
95
-9.1f
-19.4f,g
-7.2f
170
+17.8h
a 50 mg mL-1 of enzyme in diisopropyl ether, 1 equiv of H2O, 60 °C. b Yield 100% at 50% conversion. c According to GC. d Calculated by using an
internal standard (hexadecane). e c ) 0.5; CHCl3. f c ) 0.5; H2O. g [R]25 -19.6 (c 0.25, H2O).17b h c ) 0.4; H2O
D
PPL (porcine pancreatic lipase), CAL-A (lipase A from
Candida antarctica), Novozym 435 (lipase B from Candida
antarctica immobilized on a macroporous polyacrylic resin),
and Lipolase (lipase B from Candida antarctica, produced
by submerged fermentation of a genetically modified As-
pergillus oryzae microorganism and adsorbed on a macro-
porous resin) in diisopropyl ether was performed for the
model compound 3. Lipase PS, AK, AY, and PPL did not
exhibit any activity at 30 °C (no conversion after 46 h), while
CAL-A even catalyzed the hydrolysis of 3 at 60 °C, but the
reaction rate was low (Table 1, entry 15). Besides Chirazyme
L-2 (Table 1, entry 5), Lipolase (Table 1, entry 8) and
Novozym 435 (Table 1, entry 14) were all promising
catalysts, directing the hydrolysis with the same high
enantioselectivity. With regard to the reaction rates (Table
1, entries 5, 8, 14, and 15), Lipolase was chosen as the
enzyme for further studies and for the gram-scale resolution.
Several solvents were tested to study the solvent effect in
the Lipolase-catalyzed hydrolysis of (()-3 at 60 °C. Lipolase
was practically inactive in acetone and tetrahydrofuran
(conversion < 5% after 42 h). The reaction proceeded
somewhat more slowly in n-hexane (conversion 47% after
22 h) and much more slowly in toluene (conversion 37%
after 41 h) than that in diisopropyl ether (conversion 48%
after 20 h), which all afforded high enantioselectivities (E
> 68).
1 equiv of H2O, 75 mg mL-1 of Lipolase: conv. ) 50%,
ees ) 94%, eep > 95%).
On the basis of the preliminary results, the gram-scale
resolutions of (()-1-(()-4 were performed with 1 equiv of
water in the presence of Lipolase in diisopropyl ether at 60
°C. Despite the long reaction times (very different reaction
rates, probably due to the different conformations of the
cycloalkane rings), the products were characterized by an
excellent enantiomeric excess at 50% conversion. The results
are reported in Table 2 and the Supporting Information.
The transformations involving the ring opening of â-
lactams 1b-4b with 18% aqueous HCl resulted in the
enantiomers of the â-amino acid hydrochlorides 1c‚HCl-
4c‚HCl (Scheme 2). Treatment of amino acids 1a-4a with
22% HCl/EtOH resulted in enantiopure hydrochloride salts
1a‚HCl-4a‚HCl.
Scheme 2. Ring Opening of â-Lactam Enantiomers 1b-4b
with Aqueous HCl
The presence of water in the reaction medium affects the
enzymatic activity;20 accordingly, the enzyme-catalyzed ring
opening of (()-3 was performed with an increased amount
of water (Table 1, entries 5-8 and 10-13). The catalytic
activities of the tested Chirazyme L-2 and Lipolase were
progressively lowered on increase of the amount of water,
though the enantioselectivity was apparently not affected.
Furthermore, the degree of hydrolysis of 3 in the presence
of Lipolase was complete even without the addition of any
water (Table 1, entry 9), but the water in the reaction medium
(<0.1%) or present in the enzyme preparation (<5%) was
sufficient for the lactam ring opening.
Even though the E value in the Lipolase-catalyzed ring
opening is excellent, the reaction rate for the ring opening
in the case of 3 clearly increases with increasing amount of
enzyme (after 23 h, 1 equiv of H2O, 20 mg mL-1 of
Lipolase: conv ) 42%, ees ) 69%, eep > 95%; after 23 h,
The absolute configurations in the cases of 1 and 2 were
proved by comparing the [R] values with the literature data.
The value of [R]25 -5.1 (c 0.5, H2O) for 1a‚HCl and the
D
literature value21 for (1R,2S)-2-aminocyclopentanecarboxylic
acid hydrochloride, [R]25D -4.5 (c 0.5, H2O), and the value
of [R]25D -19.4 (c 0.5, H2O) for 2a and the literature value17b
for (1R,2S)-2-aminocyclohexanecarboxylic acid, [R]25D -19.6
(c 0.25, H2O), indicate (1R,5S) selectivity for the enzymatic
ring opening of (()-1 and (1R,6S) selectivity for the
enzymatic ring opening of (()-2. When (1S,8R)-9-azabicyclo-
[6.2.0]dec-4-en-10-one, recently prepared by our group,17d
was reduced catalytically in the presence of cyclohexene as
hydrogen donor, (1S,8R)-9-azabicyclo[6.2.0]decan-10-one
was formed with [R]25D -16.3 (c 0.3, CHCl3). This value is
close to the [R]25 -19 (c 0.5, CHCl3) reported for 4b.
D
Therefore, the absolute configuration for 4b is (1S,8R) and
that for 4a is (1R,2S).
(20) Brunet, C.; Zarevucka, M.; Wimmer, Z.; Legoy, M. D. Enzyme
Microb. Technol. 2002, 31, 609-614.
(21) Kanerva, L. T.; Csomo´s, P.; Sundholm, O.; Berna´th, G.; Fu¨lo¨p, F.
Tetrahedron: Asymmetry 1996, 7, 1705-1716.
Org. Lett., Vol. 5, No. 8, 2003
1211

The gas chromatograms indicate that the corresponding
enantiomer of compound 3 reacts similarly as in the cases
of 1, 2, and 4. Additionally, it was proved that the ring
opening follows the same selectivity for the five-, six-, and
eight-membered systems 1, 2, and 4, and it is therefore
doubtless that the seven-membered 3 should similarly fit the
active pocket of the enzyme. The (1S,7R) configuration is
therefore accepted for the unreactive â-lactam 3b, and
(1R,2S) for â-amino acid 4a.
In conclusion, a simple and efficient direct method was
developed for the enantioselective ring opening of unacti-
vated â-lactams 1-4 in an organic medium. The Lipolase-
catalyzed highly enantioselective reactions (E > 200), when
H2O (1 equiv) is used in diisopropyl ether at 60 °C, produce
â-amino acids (1R,2S)-1a-4a and â-lactam enantiomers
(1S,5R)-1b, (1S,6R)-2b, (1S,7R)-3b, and (1S,8R)-4b, respec-
tively, in good chemical yield (36-47%). The products could
be easily separated. Transformations by ring opening of
â-lactams 1b-4b with 18% aqueous HCl resulted in enan-
tiomers of â-amino acid hydrochlorides 1c‚HCl-4c‚HCl (ee
g 99%). The present method of formation of (1R,2S)-2-
aminocyclopentanecarboxylic acid (cispentacin) proved to
be a very simple, inexpensive route, and could be easily
scaled up.
Acknowledgment. The authors acknowledge receipt of
OTKA grants T 034901 and TS 040888, FKFP grant 0115/
2001, and a Bolyai Fellowship for E.F. (grant No.181/2002).
Supporting Information Available: Experimental de-
tails. This material is available free of charge via the Internet
at http://pubs.acs.org.
OL034096O
1212
Org. Lett., Vol. 5, No. 8, 2003