Homocarbonates as substrates for the enantioselective enzymatic protection of amines

Full text of DOI:10.1021/ja9533048

712
J. Am. Chem. Soc. 1996, 118, 712-713
Scheme 1
Homocarbonates as Substrates for the
Enantioselective Enzymatic Protection of Amines
Bernard Orsat,† Phil B. Alper,† Wilna Moree,†
Ching-Pong Mak,‡ and Chi-Huey Wong*,†
Department of Chemistry, The Scripps Research
Institute, 10666 North Torrey Pines Road
La Jolla, California 92037
Chemical Process Research and DeVelopment
Sandoz Ltd., Basle, Switzerland
ReceiVed September 28, 1995
The irreversible enzymatic acylation of alcohols in high
concentrations of organic solvents using lipases or serine
proteases as catalysts and activated esters such as choloroethyl,
trifluoroethyl, cyanomethyl, enol, oxime, or thio esters or
anhydrides as acylating reagents has been shown to be useful
in organic synthesis.1 Of these reagents, enol esters are the most
widely used,1,2 as the rate of enzymatic acylation is relatively
fast and the released enol is spontaneously tautomerized to a
ketone, making the process irreversible and free of inhibition
caused by the leaving alcohol and the product easy to isolate.
These acylating reagents, however, cannot be used in the
enzymatic acylation of amines, as they are too reactive and give
high background reactions. Recently, the use of vinyl carbon-
ates to protect amines using the lipase from Candida antarctica
has been reported.3 These unsymmetrical carbonates are,
however, not readily available and may be too reactive to avoid
nonenzymatic reactions.
Scheme 2a
We report here a novel enzymatic method for protecting
amines as carbamates with high enantioselectivity using com-
mercially available and low-cost homocarbonates as substrates
for lipases and proteases (Scheme 1).4 The reactions are
irreversible, as the products carbamates are not substrates for
the serine-type esterases. Furthermore, the symmetrical structure
of the homocarbonates gives unambiguously a single carbamate
product, making the process very simple. The carbamate can
be easily deprotected or converted to the N-methyl derivative
by reaction with LiAlH4, providing a new procedure for the
chemoenzymatic methylation of amines.
In a preliminary experiment, racemic aziridine 1 was stirred
at ambient temperature with diallyl carbonate in phosphate
buffer (pH 8.0) containing subtilisin BPN′.5 Carbamate 4 was
obtained in 50% yield after 2 h. Both dimethyl and diethyl
carbonates reacted more slowly; after 90 h, 2 and 3 were
obtained in 15 and 24% yield, respectively.6 Disappointingly,
a Conditions: (a) Homocarbonate (1 mL), CCl (20 mg), room
temperature, 45 h (0.1 mmol of 1 per milliliter of homocarbonate was
used).
the enantiomeric excess (ee) of each product, determined by
HPLC using a chiral column (Chiralcel OD-H, Daicel), was
<25%. Adding 75% 1,4-dioxane to the diallyl carbonate system
gave a maximum yield of 39% after 74 h and 54% ee.6
Switching from subtilisin BPN′ to Candida cylindracea lipase
(CCL, also called Candida rugosa lipase, CRL) and using the
carbonates as solvents at room temperature for 45 h gave better
results (Scheme 2).5,6 Due to its crystalline nature, dibenzyl
carbonate could not be used in this procedure. Racemic 4 was
also prepared by a standard procedure and used as a reference
for spectroscopic and chromatographic analyses.7
When primary amines were mixed with diallyl carbonate as
solvent, a significant background reaction was observed without
enzyme. In aqueous buffer, however, no background reaction
was detected, and the use of subtilisin BPN′ appeared to be
useful for the enzymatic protection of amines under these
conditions. To investigate the chemo- and enantioselectivity
of this process, we chose three multifunctional substrates that
could undergo different enzymatic reactions. When racemic 5
was used as substrate, the reaction was carried out in phosphate
buffer (pH 8.0) with subtilisin BPN′ and diallyl carbonate
(Scheme 3). After 69 h, 6 was obtained in 49% yield and 78%
ee, as determined by HPLC (Chiralpak AD, Daicel), based on
the O-(p-anisoyl) derivative 7 (Scheme 3).6 Carbamate 6 was
subsequently deprotected according to a standard procedure to
provide 5 after enzymatic resolution.8 The optical rotation of
the obtained 5 ([R]D +14°, c 0.8, H2O) was then compared with
the published value to establish the absolute configuration
† The Scripps Research Institute.
‡ Sandoz Ltd.
(1) For a comprehensive documentation of enzymatic acylations, see:
Klibanov, A.M. Acc. Chem. Res. 1990, 23, 114. Wong, C.-H.; Whitesides,
G. M. Enzymes in Synthetic Organic Chemistry; Pergamon: Oxford, 1994;
p 72. Fang, J.-M.; Wong, C.-H. Synlett 1994, 6, 393.
(2) Sweers, H. M.; Wong, C.-H. J. Am. Chem. Soc. 1986, 108, 6421.
Degueil-Castaing, M.; Jeso, B. D.; Drouillard, S.; Maillard, B. Tetrahedron
Lett. 1987, 28, 953. Wang, Y. F.; Lalonde, J. J.; Homongan, M.; Bergbreiter,
D. E.; Wong, C.-H. J. Am. Chem. Soc. 1988, 110, 7200. Laumen, K.;
Brietgoff, D.; Schneider, M.-P. J. Chem. Soc., Chem. Commun. 1988, 1459.
(3) Pozo, M.; Pulido, R.; Gotor, V. Tetrahedron 1992, 48, 6477. Pozo,
M.; Gotor, V. Tetrahedron 1993, 49, 10725. For other enzymatic resolutions
of amines via reactions with esters to form amides, see: Kitaguchi, H.;
Fitzpatrick, P. A.; Huber, J. E.; Klibanov, A. M. J. Am. Chem. Soc. 1989,
111, 3094. Fernandez, S.; Brieva, R.; Rebolledo, F.; Gotor, V. J. Chem.
Soc., Chem. Commun. 1992, 2885. Gotor, V.; Menendez, E.; Mouloungui,
Z.; Gasset, A. J. Chem. Soc., Chem. Commun. 1993, 2453.
(4) Dimethyl , diethyl, and diallyl carbonates were purchased from
Aldrich Chemical Co. Dibenzyl carbonate was from Lancaster Synthesis
Inc.
(5) Subtilisin BPN′ (Nagarse, type XXVII, 8.3 units/mg, P4789) and
lipase from Candida cylindracea (CCL, type VII, 860 units/mg, L1754,
EC 3.1.1.3) were purchased from Sigma.
(6) An identical reaction without enzyme did not give any detectable
product over the same period.
(7) Sennyey, G.; Barcelo, G.; Senet, J.-P. Tetrahedron Lett. 1987, 28,
5809.
(8) Kunz, H.; Unverzagt, C. Angew. Chem., Int. Ed. Engl. 1984, 23, 436.
0002-7863/96/1518-0712$12.00/0 © 1996 American Chemical Society

Communications to the Editor
J. Am. Chem. Soc., Vol. 118, No. 3, 1996 713
Scheme 3a
Scheme 4a
a (a) Diallyl carbonate (1.5 mmol, 1.5 equiv), phosphate buffer (0.1
M, pH 8.0, 10 mL), BPN′ (50 mg), room temperature, 92 h, 6% yield,
93% ee. (b) LiAlH4 (8.7 equiv), ether, 0 °C, 2 h, 93%.
b
The process was also applied to the resolution of racemic
secondary amine 12. The enzymatic reaction was carried out
for 92 h to yield 13 with 93% ee, as determined by HPLC
(Chiralcel OD-H, Daicel), and in 6% yield (Scheme 4).6 The
low yield was attributed to the steric hindrance and unfavorable
electronic effects occurring at the reacting center. The reaction
was also attempted using diallyl carbonate as solvent and CCL
as catalyst, but no reaction was observed. The carbamate was
subsequently reduced to the N-methyl derivative using LiAlH4
in ether to afford 14 in 93% yield. Resolution of racemic amino
acids can also be carried out similarly with diallyl carbonate.
Both serine and alanine, for example, were converted to the
carbamates in ∼45% yield and >95% ee.
c
a Conditions: (a) Diallyl carbonate (0.6 mmol, 3 equiv), phosphate
buffer (0.1 M, pH 8.0, 1 mL), BPN′ (10 mg), room temperature, 69 h,
49% yield, 78% ee. (b) Diallyl carbonate (0.2 mmol, 3.3 equiv),
phosphate buffer (0.1 M, pH 8.0, 1 mL), BPN′ (6 mg), room
temperature, 85 h, 45% yield, 93% ee. (c) Diallyl carbonate (1.23 mmol,
4 equiv × 4), HEPES buffer (0.2 M HEPES, 0.02 M CaCl2, pH 7.8, 3
mL), DMF (3 mL), BPN′ (10 mg), room temperature, 1 week, 76%
yield, >99% ee.
In summary, this report describes a novel enzymatic process
using commercially available, low-cost homocarbonates for the
protection of amines with high chemo- and enantioselectivity.
Diallyl carbonate was shown to be the most useful substrate
for the reaction, providing the highest yield and enantioselec-
tivity. This versatile process can be carried out in aqueous (for
primary and secondary amines) or organic solution (for second-
ary amines). Further studies are in progress to scale up some
of these reactions and to find new substrates.
(1S,2S) for the enzyme product.9 When dibenzyl carbonate was
used instead of diallyl carbonate, only a trace of product was
observed. This carbonate remained present in the reaction
mixture, indicating that it is a poor substrate for the enzyme.
Racemic â-amino ester 8 was treated similarly for 85 h, and
carbamate 9 was obtained with 93% ee, as determined by HPLC
(Chiralpak AD, Daicel), and in 45% yield without hydrolysis
of the ester (Scheme 3).6 When the concentration of 8 was
increased to 0.1 M in the presence of 11 mg/mL subtilisin, 9
was obtained in 40% yield. At 0.3 M concentration of the
substrate, 9 was obtained in 35% yield with use of 31 mg/mL
of subtilisin. The meso substrate 2-deoxystreptamine (10) was
converted to 11 through the use of subtilisin BPN′ and diallyl
carbonate in HEPES buffer (200 mM, pH 7.8, 20 mM CaCl2,
50% DMF). The chemo- and enantioselectivity of the reaction
were complete despite five possible sites of reaction. The
product was obtained in 76% yield and >99% ee, as determined
by HPLC (Chiralpak AD, Daicel), based on the peracetylated
p-toluenesulfonamide derivative 11a (Scheme 3).6
Acknowledgment. We thank the NSF for supporting the work and
the Swiss National Science Foundation for a fellowship to B.O. P.B.A.
would like to thank Pam Sears for helpful discussion.
Supporting Information Available: Experimental details for the
1
preparation of 2, 3, 4, 6, 7, 9, 11, 11a, 13, and 14 and H and 13C
NMR spectra (11 pages). This material is contained in many libraries
on microfiche, immediately follows this article in the microfilm version
of this journal, can be ordered from the ACS, and can be downloaded
from the Internet; see any current masthead page for ordering
information and Internet access instructions.
(9) Faber, K.; Ho¨nig, H.; Seufer-Wasserthal, P. Tetrahedron Lett. 1988,
29, 1903.
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