Chemoenzymatic Synthesis of Both Enantiomers of cis-6-(Hydroxymethyl)- and cis,cis-4-Hydroxy-6-(hydroxymethyl)pipecolic Acids

Article abstract of DOI:10.1021/jo9823036

Both enantiomers of cis-6-(hydroxymethyl)- and cis,cis-4-hydroxy-6-(hydroxymethyl)pipecolic acids, piperidine-based nonproteinogenic amino acids, have been synthesized from starting materials obtained from enzymatic desymmetrizations.

Full text of DOI:10.1021/jo9823036

3178
J . Org. Chem. 1999, 64, 3178-3180
Ch em oen zym a tic Syn th esis of Both En a n tiom er s of
cis-6-(Hyd r oxym eth yl)- a n d
cis,cis-4-Hyd r oxy-6-(h yd r oxym eth yl)p ip ecolic Acid s
Robert Cheˆnevert* and Marie-Pascale Morin
De´partement de chimie, Faculte´ des sciences et de ge´nie, Universite´ Laval,
Que´bec (Que´bec), Canada G1K 7P4
Received November 23, 1998
Both enantiomers of cis-6-(hydroxymethyl)- and cis,cis-4-hydroxy-6-(hydroxymethyl)pipecolic acids,
piperidine-based nonproteinogenic amino acids, have been synthesized from starting materials
obtained from enzymatic desymmetrizations.
Sch em e 1
In tr od u ction
Peptides of potential therapeutic value are easily
degraded by proteases, and their mimetics can have
superior efficacy, bioavailability, and stability. An im-
portant part of the design and synthesis of peptidomi-
metics is the availability of enantiopure, nonstandard
amino acids. They can be used to replace certain natural
amino acids in the original peptide in order to create a
peptidomimetic.1 Also, the incorporation of nonstandard
amino acids with well-defined stereochemical and struc-
tural properties is a useful tool to study peptide confor-
mation and protein folding.2 More recently, biosynthetic
methods have been developed for the site-specific incor-
poration of unnatural amino acids into proteins.3 This
expansion of the genetic code is a powerful means to
probe protein structure and function. This importance
is reflected in the number of recent original and review
publications in the literature concerning the enantiose-
lective synthesis of amino acids.4 We report here the
synthesis of both enantiomers of cis-6-(hydroxymethyl)-
pipecolic acid (4a ) and cis,cis-4-hydroxy-6-(hydroxymeth-
yl)pipecolic acid (4b) starting from alcohols 2a ,b, syn-
thons obtained by an enzymatic desymmetrization
procedure we have recently developed.5,6
Sch em e 2a
enzymatic desymmetrization of diesters 1a ,b in the
presence of Aspergillus niger lipase5 (Scheme 1). Alcohols
2a ,b were oxidized7 with RuCl3 and NaIO4 in CH3CN-
CCl4-H2O to give carboxylic acids 3a ,b (Scheme 2). Acids
3a ,b were hydrogenated (10% Pd on carbon) in acidic
water, which not only freed the amino group but also
hydrolyzed the acetate (and the MOM protecting group
in the case of 3b), to afford the amino acid hydrochlorides,
Resu lts a n d Discu ssion
Alcohols 2a ,b (ee g 98%), of known absolute configu-
ration (2a : 2R,6S; 2b: 2R,4S,6S), were obtained by
(2S,6R)-4a ([R]23 -33.6 (c 1.14, H2O)) and (2S,4R,6R)-
D
4b ([R]23 -18.2 (c 1.22, H2O)).
(1) (a) Ripka, A. S.; Rich, D. H. Curr. Opin. Chem. Biol. 1998, 2,
441. (b) Giannis, A.; Rubsam, F., Adv. Drug Res. 1997, 29, 1. (c)
Liskamp, R. M. J . Angew. Chem., Int. Ed. Engl. 1994, 33, 305. (d)
Gante, J . Angew. Chem., Int. Ed. Engl. 1994, 33, 1699. (e) Symposia-
in-Print No. 5, Horwell, D. C. (Guest Editor). Bioorg. Med. Chem. Lett.
1993, 3, 797.
(2) (a) Filigheddu, S. N.; Taddei, M. Tetrahedron Lett. 1998, 39,
3857. (b) Kahn, M. Synlett 1993, 821. (c) Symposia-in-Print No. 50,
Kahn, M. (Guest Editor). Tetrahedron 1993, 49, 3433.
D
The opposite enantiomers were synthesized from the
same starting materials 2a ,b (Scheme 3). Protection of
the 6-hydroxy group of 2a ,b with chloromethyl methyl
ether (MOM) in the presence of diisopropylethylamine
followed by the hydrolysis of the acetates 5a ,b with pig
liver esterase (PLE) gave 6a ,b in high yields. The use of
an hydrolase was necessary because base-catalyzed hy-
drolysis gave an oxazolidinone from the attack of the
alcoholate on the N-benzyloxycarbonyl group. Oxidation
of 6a ,b by the method mentioned above provided the
carboxylic acids 7a ,b which were hydrogenated in an
acidic aqueous solution, thus provoking the removal of
the benzyl carbamate as well as the MOM groups to
(3) Cornish, V. W.; Mendel, D.; Schultz, P. G. Angew. Chem., Int.
Ed. Engl. 1995, 34, 621.
(4) (a) Hanessian, S.; McNaughton-Smith, G.; Lombart, H. G.;
Lubell, W. D. Tetrahedron 1997, 53, 12789. (b) Sardina, F. J .; Rapoport,
H. Chem. Rev. 1996, 96, 1825. (c) Duthaler, R. O. Tetrahedron 1994,
50, 1539. (d) Ohfune, Y. Acc. Chem. Res. 1992, 25, 360. (e) Williams,
R. M. Organic Chemistry Series: Synthesis of Optically Active R-Amino
Acids; Baldwin, J . E., Magnus, P. D., Eds.; Pergamon: Oxford, 1989.
(f) Barrett, G. C. Chemistry and Biochemistry of the Amino Acids;
Chapman and Hall, London, 1985.
(5) Cheˆnevert, R.; Dickman, M. J . Org. Chem. 1996, 61, 3332.
(6) For a preliminary report, see: Cheˆnevert, R.; Morin, M. P.
Tetrahedron: Asymmetry 1996, 7, 2161.
(7) Carlsen, P. H. J .; Katsuki, T.; Martin, V. S.; Sharpless, K. B. J .
Org. Chem. 1981, 46, 3936.
10.1021/jo9823036 CCC: $18.00 © 1999 American Chemical Society
Published on Web 04/03/1999

Chemoenzymatic Synthesis of Enantiomers
J . Org. Chem., Vol. 64, No. 9, 1999 3179
CHCl3); IR (neat) 3025, 2925, 1745, 1695 cm-1 1H NMR
Sch em e 3a
;
(CDCl3) 7.35-7.23 (m, 5H), 5.12 (AB system, J ) 12.4 Hz, 2H),
4.70 (s, 2H), 4.55-4.36 (m, 2H), 4.13 (dd, J 1 ) 10.5 Hz, J 2
8.4 Hz, 1H), 3.85 (dd, J 1 ) 10.6 Hz, J 2 ) 6.4 Hz, 1H), 3.52-
3.39 (m, 2H), 3.28 (s, 3H), 1.94 (s, 3H), 1.86-1.45 (m, 6H); 13
)
C
NMR (CDCl3) 170.5, 155.9, 136.5, 128.3, 127.8, 127.7, 96.2,
67.9, 67.1, 64.1, 55.1, 49.3, 48.2, 24.8, 24.7, 20.6, 14.2; HRMS
(EI) calcd for C19H27NO6 (M+) 366.1916 found 366.1920 (
0.0011.
(-)-N-(Ben zyloxycar bon yl)-cis,cis-2(R)-(acetoxym eth yl)-
4(S)-(m eth oxym eth oxy)-6(S)-[(m eth oxym eth oxy)m eth yl)]-
p ip er id in e (5b). Flash chromatography with 40% EtOAc/60%
petroleum ether afforded 5b in 95% yield as a colorless oil.
[R]25D ) -6.15 (c 2.02, CHCl3); IR (neat) 3025, 2910, 1740, 1695
cm-1; 1H NMR (CDCl3) 7.37-7.27 (m, 5H), 5.13 (AB system, J
) 12.4 Hz, 2H), 4.66-4.53 (m, 5H), 4.45-4.42 (m, 1H), 4.29
(dd, J 1 ) 7.0 Hz, J 2 ) 10.7 Hz, 1H), 4.11 (dd, J 1 ) 7.7 Hz, J 2
) 10.7 Hz, 1H), 3.96-3.92 (m, 1H), 3.75 (t, J ) 9.0 Hz, 1H),
3.60 (dd, J 1 ) 5.6 Hz, J 2 ) 9.2 Hz, 1H), 3.33 (s, 3H), 3.31 (s,
3H), 1.92 (s, 3H), 1.87-1.74 (m, 4H); 13C NMR (CDCl3) 170.5,
155.9, 136.5, 128.3, 127.9, 127.8, 96.3, 94.2, 69.7, 67.9, 67.2,
66.3, 55.3, 55.1, 48.8, 47.6, 29.2, 28.5, 20.6; HRMS (EI) calcd
for C21H31NO8 (M+) 426.2128, found 426.2133 ( 0.0013.
Gen er a l P r oced u r e for En zym a tic Hyd r olysis of Ac-
eta tes 5a a n d 5b. The acetate (0.8 mmol) was suspended in
20 mL of phosphate buffer at pH 7.0. To this mixture was
added PLE (100 mg), and the reaction mixture was stirred at
room temperature. The pH of the solution was maintained at
its initial value by addition of 0.1 N NaOH. After the addition
of 1 equiv of base (∼24 h), the aqueous mixture was saturated
with NaCl and extracted three times with EtOAc. The com-
bined EtOAc fractions were dried (MgSO4) and evaporated.
afford enantiopure amino acid hydrochlorides, (2R,6S)-
4a ([R]23 +34.5 (c 1.14, H2O)) and (2R,4S,6S)-4b ([R]23
D
D
+17.4 (c 1.32, H2O)).
Both enantiomers of substituted pipecolic acids 4a ,b
have been prepared from the same starting materials
2a ,b, which were obtained by enzymatic desymmetriza-
tion of meso compounds 1a ,b. These nonproteinogenic
amino acids could be incorporated into peptidomimetics
for their secondary structure-promoting effects and in-
creased proteolytic stability. Pipecolic acid derivatives
4a ,b possess a set of characteristics which makes them
attractive scaffolds8 for combinatorial synthesis: they are
enantiomerically pure and conformationally rigid; they
provide a three-dimensional spatial arrangement of
orthogonally manipulable functional groups for the gen-
eration of molecular diversity. Also, the piperidine ring
is a common structural subunit in natural products or
bioactive synthetic compounds. The interest in these
compounds is well-displayed by the wealth of published
material detailing their sources, biological activities, and
syntheses.9
(-)-N-(Ben zyloxycar bon yl)-cis-2(S)-[(m eth oxym eth oxy)-
m eth yl]-6(R)-(h yd r oxym eth yl)p ip er id in e (6a ). Flash chro-
matography with 50% EtOAc/50% petroleum ether gave 6a
in 85% yield as a colorless oil. [R]25 ) -5.7 (c 1.1, CHCl3); IR
D
(neat) 3680-3140, 3030, 2930, 1680 cm-1 1H NMR (CDCl3)
;
7.34-7.27 (m, 5H), 5.13 (AB system, J ) 12.4 Hz, 2H), 4.55-
4.52 (m, 2H), 4.41-4.33 (m, 2H), 3.56-3.44 (m, 4H), 3.27 (s,
3H), 2.92 (br s, 1H), 1.76-1.44 (m, 6H); 13C NMR (CDCl3)
156.9, 136.5, 128.3, 127.8, 127.7, 96.2, 67.9, 67.2, 64.3, 55.2,
51.7, 49.3, 24.8, 24.4, 14.5; HRMS (EI) calcd for C17H25NO5
(M+) 326.1603, found 326.1607 ( 0.0010.
(+)-N-(Ben zyloxycar bon yl)-cis,cis-2(S)-[(m eth oxym eth -
oxy)m eth yl]-4(R)-(m eth oxym eth oxy)-6(R)-(h ydr oxym eth -
yl)p ip er id in e (6b). Flash chromatography with 75% EtOAc/
25% petroleum ether gave 6b in 92% yield as a colorless oil.
[R]25D ) +5.6 (c 1.24, CHCl3); IR (neat) 3620-3120, 3020, 2930,
Exp er im en ta l Section
Gen er a l. NMR spectra were recorded at 300 MHz (1H) and
75 MHz (13C). Melting points are uncorrected. Flash column
chromatography was carried out using 230-400 mesh silica
gel. Pig liver esterase was from Sigma or Amano.
Gen er a l P r oced u r e for O-P r otection of Alcoh ols 2a
a n d 2b. To a solution of 2a or 2b (6 mmol) and 3.5 equiv of
diisopropyl ethylamine in 20 mL of anhydrous CH2Cl2 at 0 °C
under N2 were added 3 equiv of MOM-Cl dropwise with
stirring. After 30 min, the ice bath was removed, and the
reaction was stirred for 15 h. The reaction mixture was diluted
with 200 mL of EtOAc and washed successively with 1 N HCl
(3 × 25 mL), saturated aqueous NaHCO3 (3 × 25 mL), and
brine (50 mL). The organic phase was dried (MgSO4) and
evaporated.
1690 cm-1 1H NMR (CDCl3) 7.33-7.26 (m, 5H), 5.13 (AB
;
system, J ) 12.3 Hz, 2H), 4.62-4.54 (m, 4H), 4.47-4.41 (m,
2H), 3.90-3.84 (m, 1H), 3.79-3.61 (m, 4H), 3.33 (s, 3H), 3.28
(s, 3H), 3.01 (br s, 1H), 1.96-1.80 (m, 4H); 13C NMR (CDCl3)
156.6, 136.4, 128.3, 127.9, 127.8, 96.1, 94.7, 79.8, 69.6, 68.8,
67.3, 65.6, 55.3, 55.2, 51.5, 49.2, 29.1; HRMS (EI) calcd for
C
19H29NO7 (M+) 384.2022, found 384.2018 ( 0.0011.
Gen er a l P r oced u r e for Oxid a tion of Alcoh ols 2a , 2b,
6a , 6b. The alcohol (0.6 mmol) was dissolved into a mixture
of acetonitrile (1.2 mL), CCl4 (1.2 mL), and water (1.8 mL).
RuCl3‚3H2O (0.022 equiv) and NaIO4 (4.1 equiv) were succes-
sively added, and the mixture was stirred overnight at room
temperature. Brine (10 mL) was added to the mixture, and
the aqueous phase was extracted four times with CH2Cl2. The
combined organic phases were dried (MgSO4) and evaporated.
The crude product was purified by flash chromatography with
silica gel neutralized with Et3N (3% w/w).
(-)-N-(Ben zyloxyca r bon yl)-cis-2(R)-(a cetoxym eth yl)-
6(S)-[(m et h oxym et h oxy)m et h yl]p ip er id in e (5a ). Flash
chromatography with 25% EtOAc/75% petroleum ether af-
forded 5a in 94% yield as a colorless oil. [R]25 ) -11.6 (c 1.0,
D
(-)-N-(Ben zyloxyca r bon yl)-cis-6(R)-(a cetoxym eth yl)-
p ip er id in e-2(S)-ca r boxylic Acid (3a ). Flash chromatogra-
phy with CHCl3-EtOAc-MeOH (14:1:1) gave acid 3a in 73%
(8) (a) Pavia, M. R. Combinatorial Chemistry and Molecular Diver-
sity in Drug Discovery; Gordon, E. M., Kerwin, J . F., Eds.; Wiley: New
York, 1998; Chapter 11, pp 213-225. (b) Sofia, M. J .; Hunter, R.; Chan,
T. Y.; Vaughan, A.; Dulina, R.; Wang, H.; Gange, D. J . Org. Chem.
1998, 63, 2802.
(9) (a) Bailey, P. D.; Millwood, P. A.; Smith, P. D. Chem. Commun.
1998, 633. (b) Wang, C. L. J .; Wuonola, M. A. Org. Prep. Proc. Int.
1992, 24, 583. (c) Barluenga, J .; Aznar, F.; Valde´s, C.; Ribas, C. J .
Org. Chem. 1998, 63, 3918.
yield as a colorless oil. [R]25 ) -17.7 (c 1.0, CHCl3); IR (neat)
D
3640-3310, 3030, 2930, 1745, 1705 cm-1 1H NMR (CDCl3)
;
10.35 (br s, 1H), 7.34-7.26 (m, 5H), 5.19 (m, 2H), 4.93-4.84
(m, 1H), 4.46 (s, 1H), 4.17 (m, 2H), 2.32 (m, 1H), 1.98 (s, 3H),
1.75-1.56 (m, 5H); 13C NMR (CDCl3) 176.3, 170.8, 156.4, 136.7,

3180 J . Org. Chem., Vol. 64, No. 9, 1999
Cheˆnevert and Morin
128.2, 127.6, 127.5, 67.0, 63.3, 54.1, 49.1, 26.4, 24.8, 20.8, 15.8;
HRMS (EI) calcd for C17H21NO6 (M+) 336.1447, found 336.1451
( 0.0010.
Gen er a l P r oced u r e for Hyd r ogen a tion of 3a , 3b, 7a ,
a n d 7b. To a suspension of the amine (0.25 mmol) in 6 N HCl
(25 mL) was added 75 mg of Pd/C, and the mixture was
hydrogenated at 45 psi at 50 °C for 15 h. After filtration of
the mixture through a pad of Celite, water was evaporated.
Column chromatography on Sephadex G-10 (40-120 µm) with
MeOH as eluant gave the amino acid as a white solid. The
product was recrystallized in H2O-EtOH.
(+)-N-(Ben zyloxyca r bon yl)-cis,cis-4(R)-(m eth oxym eth -
oxy)-6(R)-(acetoxym eth yl)piper idin e-2(S)-car boxylic Acid
(3b). Flash chromatography with CHCl3-EtOAc-MeOH (10:
1:1) gave acid 3b in 75% yield as a colorless oil. [R]25D ) +28.1
(c 2.4, CHCl3); IR (neat) 3650-3300, 3035, 3030, 1750, 1705
cm-1; 1H NMR (CDCl3) 7.78 (br s, 1H), 7.27-7.19 (m, 5H), 5.09
(s, 2H), 4.73 (s, 1H), 4.55-4.53 (m, 1H), 4.42-4.40 (m, 2H),
4.32 (s, 2H), 3.91 (s, 1H), 3.26 (s, 3H), 2.57-2.53 (m, 1H), 1.87
(s, 3H), 1.75-1.63 (m, 3H); 13C NMR (CDCl3) 176.8, 170.8,
156.4, 136.1, 128.4, 127.9, 127.8, 93.7, 67.7, 65.9, 65.8, 55.2,
49.8, 48.0, 29.1, 28.7, 20.7; HRMS (EI) calcd for C19H25NO8
(M+) 396.1658, found 396.1666 ( 0.0012.
cis-6-(Hydr oxym eth yl)piper idin e-2-car boxylic Acid (4a).
(2S,6R)-4a : [R]25 ) -33.6 (c 1.14, H2O); (2R,6S)-4a : [R]25
)
D
D
+34.5 (c 1.14, H2O). Spectroscopic data are identical for both
enantiomers: mp 225-230 °C (dec); IR (KBr) 3400, 3250-2500,
1780 cm-1; 1H NMR (D2O) 3.89 (dd, J 1 ) 12.3 Hz, J 2 ) 3.3 Hz,
1H), 3.77 (dd, J 1 ) 12.6 Hz, J 2 ) 3.8 Hz, 1H), 3.60 (dd, J 1
)
12.6 Hz, J 2 ) 7.2 Hz, 1H), 3.24 (m, 1H), 2.26 (m, 1H), 1.88 (m,
2H), 1.62 (m, 2H), 1.41 (m, 1H); 13C NMR (D2O) 174.2, 64.2,
60.9, 60.2, 28.3, 26.4, 24.1; HRMS (EI) calcd for C7H14ClNO3
(M+) 160.0974, found 160.0976 ( 0.0005.
(+)-N-(Ben zyloxycar bon yl)-cis-6(S)-[(m eth oxym eth oxy)-
m eth yl]p ip er id in e-2(R)-ca r boxylic Acid (7a ). Flash chro-
matography with CHCl3-EtOAc-MeOH (14:1:1) gave acid 7a
cis,cis-4-Hyd r oxy-6-(h yd r oxym eth yl)p ip er id in e-2-ca r -
in 81% yield as a colorless oil. [R]25 ) +15.4 (c 1.6, CHCl3);
D
boxylic Acid (4b). (2S,4R,6R)-4b: [R]25D ) -18.2 (c 1.22, H2O);
IR (neat) 3450, 3060, 2940, 1730, 1700 cm-1; 1H NMR (CDCl3)
10.89 (br s, 1H), 7.32-7.26 (m, 5H), 5.17 (AB system, J ) 11.1
Hz, 2H), 4.95-4.88 (m, 1H), 4.58-4.48 (m, 2H), 4.35 (m, 1H),
3.68-3.45 (m, 2H), 3.28 (d, J ) 16.9 Hz, 3H), 2.31 (m, 1H),
1.93 (m, 1H), 1.69-1.53 (m, 4H); 13C NMR (CDCl3) 176.3,
156.8, 136.2, 128.3, 127.9, 127.8, 96.1, 67.7, 66.7, 54.9, 52.9,
50.1, 25.9, 24.5, 15.6; HRMS (EI) calcd for C17H23NO6 (M+)
338.1603, found 338.1608 ( 0.0010.
(2R,4S,6S)-4b : [R]25 ) +17.02 (c 1.32, H2O). Spectroscopic
D
data are identical for both enantiomers: mp 205-208 °C (dec);
IR (KBr) 3420, 3275-2520, 1770 cm-1 1H NMR (D2O) 3.97
;
(d, J ) 12.5 Hz, 2H), 3.81-3.78 (m, 1H), 3.67-3.61 (m, 1H),
3.37-3.26 (m, 1H), 2.51 (d, J ) 12.7 Hz, 1H), 2.09 (d, J ) 12.9
Hz, 1H), 1.60 (dd, J 1 ) 12.6, J 2 ) 12.1, 1H), 1.43 (dd, J 1
)
12.5 Hz, J 2 ) 12.2 Hz, 1H); 13C NMR (D2O) 174.5, 67.8, 63.9,
59.0, 36.4, 35.1; HRMS (EI) calcd for C7H14ClNO4 (M+)
176.0923, found 176.0928 ( 0.0005.
(-)-N-(Ben zyloxyca r bon yl)-cis,cis-4(S)-(m eth oxym eth -
oxy)-6(S)-[(m eth oxym eth oxy)m eth yl]piper idin e-2(R)-car -
boxylic Acid (7b). Flash chromatography with CHCl3-
EtOAc-MeOH (10:1:1) gave acid 7b in 78% yield as a colorless
Ack n ow led gm en t. The authors thank the Natural
Sciences and Engineering Research Council of Canada
(NSERC) for financial support as well as NSERC and
“le Fonds pour les chercheurs et l’aide a` la recherche,
Que´bec”, for postgraduate scholarships to M.P.M.
oil. [R]25 ) -30.8 (c 3.6, CHCl3); IR (neat) 3380, 3040, 2940,
D
1740, 1690 cm-1; H NMR (CDCl3) 9.97 (br s, 1H), 7.33-7.25
1
(m, 5H), 5.22-5.18 (m, 2H), 4.83 (s, 1H), 4.62-4.59 (m, 3H),
4.50 (d, J ) 6.9 Hz, 1H), 4.37 (s, 1H), 4.00 (m, 1H), 3.86-3.78
(m, 2H), 3.31 (s, 6H), 2.60 (d, J ) 12.4 Hz, 1H), 2.09 (d, J )
14.1 Hz, 1H), 1.87-1.72 (m, 2H); 13C NMR (CDCl3) 176.8,
156.3, 136.2, 128.4, 127.9, 127.8, 96.2, 93.6, 69.0, 67.7, 66.2,
Su p p or tin g In for m a tion Ava ila ble: Spectrometric in-
formation (1H NMR) for new compounds. This material is
available free of charge via the Internet at http://pubs.acs.org.
55.2, 55.0, 50.5, 48.9, 29.1, 28.2; HRMS (EI) calcd for C19H27
-
NO8 (M+) 398.1815, found 398.1822 ( 0.0012.
J O9823036