Synthesis of both enantiomers of hydroxypipecolic acid derivatives equivalent to 5-azapyranuronic acids and evaluation of their inhibitory activities against glycosidases

Article abstract of DOI:10.1016/j.bmc.2008.06.016

We have synthesized 3-hydroxy- and 3,4,5-trihydroxypipecolic acid derivatives corresponding to 5-aza derivatives of uronic acids and evaluated their inhibitory activities against various glycosidases including β-glucuronidase. Compounds 4 and 5 were chosen as common intermediates for the synthesis of 3,4,5-trihydroxypipecolic acids and 3-hydroxypipecolic acids as well as for 3-hydroxybaikiain, a unique natural product isolated from a toxic mushroom. Cross aldol reaction of N-Boc-allylglycine derivative with acrolein followed by the ring-closing metathesis gave 4 and 5 as a mixture of diastereomers which could be separated by silica gel column chromatography. By employing lipase-catalyzed kinetic resolution, the synthesis of both l- and d-isomers of 3,4,5-trihydroxy- and 3-hydroxypipecolic acids was achieved. None of the compounds tested showed inhibitory activity against α- and β-glucosidases. On the other hand, l-23 and l-29 were found to have potent inhibitory activity against β-glucuronidase. In addition, it is interesting that some uronic-type azasugar derivatives showed moderate inhibitory activities against β-N-acetylglucosaminidase.

Full text of DOI:10.1016/j.bmc.2008.06.016

Bioorganic & Medicinal Chemistry 16 (2008) 8273–8286
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Synthesis of both enantiomers of hydroxypipecolic acid derivatives
equivalent to 5-azapyranuronic acids and evaluation of their
inhibitory activities against glycosidases
Yuichi Yoshimura a, Chiaki Ohara a, Tatsushi Imahori a, Yukako Saito a, Atsushi Kato b,
Saori Miyauchi b, Isao Adachi b, Hiroki Takahata a,
*
a Faculty of Pharmaceutical Sciences, Tohoku Pharmaceutical University, 4-4-1, Komatsushima, Aoba-ku, Miyagi, Sendai 981-8558, Japan
b Department of Hospital Pharmacy, University of Toyama, Toyama 930-0194, Japan
a r t i c l e i n f o
a b s t r a c t
Article history:
We have synthesized 3-hydroxy- and 3,4,5-trihydroxypipecolic acid derivatives corresponding to 5-aza
derivatives of uronic acids and evaluated their inhibitory activities against various glycosidases including
b-glucuronidase. Compounds 4 and 5 were chosen as common intermediates for the synthesis of 3,4,5-
trihydroxypipecolic acids and 3-hydroxypipecolic acids as well as for 3-hydroxybaikiain, a unique natural
product isolated from a toxic mushroom. Cross aldol reaction of N-Boc-allylglycine derivative with acro-
lein followed by the ring-closing metathesis gave 4 and 5 as a mixture of diastereomers which could be
separated by silica gel column chromatography. By employing lipase-catalyzed kinetic resolution, the
Received 16 May 2008
Revised 7 June 2008
Accepted 10 June 2008
Available online 13 June 2008
Keywords:
Hydroxypipecolic acid
Glycosidase inhibitor
Azasugar
synthesis of both
of the compounds tested showed inhibitory activity against
23 and -29 were found to have potent inhibitory activity against b-glucuronidase. In addition, it is inter-
L- and
D-isomers of 3,4,5-trihydroxy- and 3-hydroxypipecolic acids was achieved. None
a
- and b-glucosidases. On the other hand,
L-
L
Uronic type
esting that some uronic-type azasugar derivatives showed moderate inhibitory activities against b-N-
acetylglucosaminidase.
Both enantiomers
Ó 2008 Elsevier Ltd. All rights reserved.
1. Introduction
OH
OH
OH
OH
HO
OH
OH
Azasugars (iminosugars) are a unique class of compounds that
have inhibitory activities against various glycosidases by acting
as transition state analogues, and they have potential applications
as drugs, including antidiabetics, antiobesities, antivirals and ther-
apeutics, for the treatment of some genetic disorders such as Gau-
cher disease.1 we have recently been interested in the
development of novel synthetic methods for azasugars and their
applications to search for a new glycosidase inhibitor as well as
to study the structural–activity relationship (SAR) between aza-
sugars and glycosidases.2 SAR studies of azasugars are important
for designing novel glycosidase inhibitors based on an azasugar
scaffold. For instance, we2a and others2b reported the syntheses
of 1-deoxynojirimycin (DNJ), fagomine, isofagomine and their ster-
eoisomers (Chart 1), and through the SAR studies of these ana-
N
H
N
H
1-Deoxynojirimycin
Fagomine
OH
HO
OH
N
H
Isofagomine
Chart 1.
of uronic acids (Chart 2). It is widely known that D-glucuronic acid,
logues, we revealed that an
acting as a non-competitive inhibitor for
As a part of our continuous synthetic study for azasugars, we
became interested in hydoroxylated pipecolic acid derivatives.
3,4,5-Trihydroxypipecolic acids are considered as 5-aza congeners
L
-isomer of DNJ was capable of
a typical uronic acid, plays a crucial role in drug metabolism
known as ‘glucuronic acid conjugation’. b-Glucuronidase is an en-
zyme that hydrolyzes a b-glycosidic bond of a terminal glucuronic
acid residue in oligo- and polysaccharides. Inhibitors of b-glucu-
ronidase are clinically important since this class of inhibitors has
recently been shown to have a protective effect against antitumor
camptotecin derivative CPT-11-induced mucosa damage and diar-
a
-glucosidase.2a
* Corresponding author. Tel./fax: +81 22 727 0144.
E-mail address: takahata@tohoku-pharm.ac.jp (H. Takahata).
rhea.3 The 5-aza analogue of
D-glucuronic acid, one of the 3,4,5-tri-
0968-0896/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2008.06.016

8274
Y. Yoshimura et al. / Bioorg. Med. Chem. 16 (2008) 8273–8286
LiHMDS (1.2 eq.)
Acrolein (1.5 eq.)
OH
OH
OH
Boc
CO2Et
N
THF
N
CO2Et
N
CO2Et
N
COOH
2
Boc
-80 oC, 1 h
87%
Boc
H
1
3
3-Hydroxybaikiain
Grubbs 1st
(1 mol%)
OH
OH
CO2Et
OH
OH
+
CH2Cl2
N
N
CO2Et
N
COOH
HO
OH
r.t., 12 h
99 %
Boc
Boc
H
4 : 5 = 4 : 1
3-Hydroxypipecolic acid
4
5
N
H
COOH
Scheme 1.
Trihydroxypipecolic acid
Chart 2.
Along with our initial plan to obtain both enantiomers of the
common intermediate 4, a kinetic resolution by using a lipase-cat-
alyzed transesterification was tried. Among the lipases tested, li-
pase PS-C (lipase isolated from Pseudomonas cepacia immobilized
on ceramic particles, purchased from Amano) gave the best result
(Scheme 2). The reaction conditions were further optimized and
the results are summarized in Table 1.
hydroxypipecolic acid derivatives, is a potential b-glucuronidase
inhibitor which may act as a transition-state analogue. However,
there have been only a few reports on glycosidase inhibition of
uronic acid-type azasugar derivatives.4
There is a strong demand for a practical route providing an easy
access to 3,4,5-trihydroxypipecolic acid derivatives, and a chiral
synthesis constitutes an area of considerable current interest. We
envisioned synthesis of 3,4,5-trihydroxypipecolic acids from a pre-
cursor 1, which would be prepared by an enzymatic reaction to ob-
tain both of the enantiomers in one synthetic scheme. Combined
with the enzymatic preparation, the conversion of a double bond
of 1 to a diol moiety with control of their stereochemistries allows
all stereoisomers of trihydroxypipecolic acids to be synthesized.
Such a synthetic strategy will be of benefit to SAR study for these
analogues. In addition, 1 is also a useful intermediate for synthesiz-
ing 3-hydroxypipecolic acids which constitute non-natural vari-
ants of a structural motif often encountered in a variety of
functional molecules and may be regarded as expanded hydroxyl-
ated proline or a conformationally restricted serine derivative.5
(ꢀ)-3-Hydroxybaikiain, the 4,5-dehydro derivative of 2,3-cis-3-hy-
When hexane and carbon tetrachloride were used as solvents,
the reaction gave
ees of an acetate
D
-6 and
L-4 in good yields, but neither of the
L
-4 was satisfactory (entries 1 and 2).11 The reac-
tion without a solvent gave a similar result (entry 3). The best re-
sult was obtained when diisopropyl ether was used, and the
reaction gave 47% of
Compound -4, a chiral precursor for
tives, in hand was converted to 2,3-trans-3-hydroxybaikiain
and 2,3-trans-3-hydroxypipecolic acid -9. Treatment of -4 with
5 N HCl at 120 °C for 3 h gave 2,3-trans-3-hydroxybaikiain (3-epi-
3-hydroxybaikiain) -7 in 99% yield after ion-exchange column
chromatography. Similarly, compound -8 obtained from hydroge-
nation of -4 was treated with refluxing 5 N HCl to give 2,3-trans-3-
hydroxypipecolic acid -9 in 84% yield (Scheme 3).
D
-6 with 97%ee and 47% of
-pipecolic acid deriva-
-7
L-6 with 99%ee.
L
L
L
L
L
L
L
L
L
Next our target was shifted to 3-hydroxybaikiain, a natural
product isolated from a toxic mushroom.6 To achieve the synthesis
of 3-hydroxybaikiain, the synthesis should be starting from the cis-
isomer 5 by using the same schemes as those described above.
Since 5 was the minor product of the sequential cross aldol and
RCM reactions, this plan did not seem efficient. Thus, we intended
droxy-L-pipecolic acid, has been isolated from a toxic mushroom,
Russula subnigricans Hongo.6 However, a simultaneous synthesis
of all stereoisomers of 3-hydroxypipecolic acid and the chiral syn-
thesis of their 4,5-dehydro compounds have not been achieved.7 To
the best of our knowledge, their inhibitory activities against glyco-
sidases have never been investigated. Herein, we report a new chi-
ral synthesis of stereoisomers of 3-hydroxy- and 3,4,5-
trihydroxypipecolic acids as well as 3-hydoxybaikiain derivatives
in conjunction with their glycosidases such as b-glucuronidase
inhibitory activities.8
LipasePS-C
Vinyl acetate (excess)
OAc
OH
4
+
solvent
temp., time
N
CO2Et
N
CO2Et
2. Chemistry
Boc
Boc
-4
N-Boc-allylglycine ethyl ester 2,9 readily obtainable from ethyl
bromoacetate and allylamine in two steps, was treated with lith-
ium hexamethyldisilazide (LiHMDS) to generate a corresponding
enolate, a reaction of which with acrolein gave an inseparable mix-
ture of 3 in good yield. Because of the presence of rotarmors caused
by the N-Boc protecting group, it was difficult to determine the ra-
tio of syn and anti adducts from its NMR spectrum. A ring-closing
metathesis (RCM) reaction of 3 afforded a mixture of 4 and 5,
key intermediates for hydroxypipecolic acids, which were sepa-
rated by silica gel column chromatography in 80% and 19%, respec-
tively (Scheme 1). Complicated NMR spectra of 4 and 5 caused by a
non-bonded interaction of N-Boc protecting groups made it diffi-
cult to elucidate their relative stereochemistries. Thus, the major
isomer 4 was converted to a known compound in three steps
and it was determined as a trans adduct (data not shown).10
D
-6
L
Scheme 2.
Table 1
Summary of lipase PS-C-catalyzed transesterification
Entry
Solvent
Temperature, time
D-6
L-4
Yielda (%)
%eeb
Yielda (%)
%eeb
1
2
3
4
Hexane
CCl4
None
i-Pr2O
40 °C, 3 days
40 °C, 3 days
30 °C, 4 days
40 °C, 3 days
44
41
45
47
95
99
92
97
54
53
52
47
87
68
70
99
a
Isolated yield.
b
The ees were determined by HPLC (see Section 5).

Y. Yoshimura et al. / Bioorg. Med. Chem. 16 (2008) 8273–8286
8275
OH
OH
5N-HCl
5N-HCl
L
-4
L
-5
120 oC, 3 h
99 %
120 oC, 5 h
85 %
N
COOH
N
COOH
H
H
H2 / Pd-C
EtOAc
r.t., 6 h
99 %
H2 / Pd-C
EtOAc
r.t., 4 h
88 %
L
-7
L
-11
OH
OH
OH
CO2Et
OH
5N-HCl
5N-HCl
120 oC, 9.5 h
86 %
N
H
COOH
COOH
N
H
120 oC, 3 h
84 %
N
CO2Et
N
Boc
-12
Boc
L
-9
L
L-13
L-8
Scheme 5.
Scheme 3.
to synthesize a chiral intermediate L-5, started from the major iso-
mer 4, by employing the Mitsunobu reaction and enzymatic
hydrolysis.
Compound 4 was treated with triphenylphsphine and acetic
acid in the presence of diethyl azodicarboxylate (DEAD) to give in-
verted acetate 10 which contains a small amount of inseparable
impurity. Lipase-mediated hydrolysis of acetate 10 gave a desired
LiOH
MeOH-H2O
OH
CO2Et
-30 oC, 10 min
D
-6
N
97 %
Boc
-4
D
chiral synthon
contained impurity, both
purities over 99%ee (Scheme 4).12
Deprotection of -5 by acid treatment gave 3-hydroxybaikiain
11 in 85% yield. All of the spectral data of -11 were identical to
those reported,6 and the structure of
-11 was therefore unambig-
uously determined. Similar to the trans isomer, -13 was synthe-
sized from -5 in two steps as shown in Scheme 5.
The -isomers of 3-hydroxypipecolic acids were the final target
of this series and were to be obtained from common intermediates
-4 and -5. Compound -4 was readily obtained from -6 by
hydrolysis of an acetyl group treated with lithium hydroxide in
97% yield. Deacetylation of -10 with LiOH was rather complicated
L
-5 in 30% yield along with
D-10. Although D-10 still
Scheme 6.
L
-5 and -10 showed high enatiomeric
D
L
L-
D
-10
L
LiOH
MeOH-H2O
L
L
-30 oC, 30 min
L
D
OH
CO2Me
OH
D
D
D
D
N
N
CO2Et
N
CO2Et
D
Boc
Boc
Boc
since a competitive b-elimination product 14 and a transesterifica-
tion product 15 were accompanied. The reaction, however, gave
D
-15
D
-5
14
12%
2%
60%
pure
D-5 in 60% yield after purification by silica gel column chro-
Scheme 7.
matography. By using the same schemes as those mentioned
above, the syntheses of
D
-enantiomers of 3-hydroxybaikiains (
-11) and 3-hydroxypipecolic acids ( -9 and -13) from
-5 were achieved (see Section 5) (Schemes 6 and 7).
D
D
-7
-4
and
and
D
D
D
tion of L-4 with catalytic potassium osmate in the presence of N-
D
methylmorpholine N-oxide (NMO) gave a diastereomeric mixture
of triol derivatives, which were separated by silica gel column
As a final series of hydroxypipecolic acids for SAR study, we
aimed to synthesize all stereoisomers of 3,4,5-trihydroxypipecolic
chromatography to give
respectively. The structures of
by converting to the corresponding nojirimycin derivatives.13 As a
L-16 and
L
-17 in 32% and 59% yields,
acids. First, cis-dihydroxylation of
L-4 by OsO4 was tried. The reac-
L-16 and
L-17 were firmly elucidated
DEAD
result, the major
17 was a 3,4-trans isomer. The resulting triols
treated with refluxing 5 N HCl to give trihydroxypipecolic acids
1814 and
-19.
In comparison with the structure of azasugar,
L
-16 had relative stereochemistry of 3,4-cis and
L
-
PPh3
L
-16 and
L-17 were
OAc
AcOH
L-
4
L
THF
r.t., 4 h
<90 %
N
CO2Et
Boc
L-18 and
L
-19 cor-
responded to 5-azaderivatives of
acids, respectively (Scheme 8).
D-allo- and D-mannopyranuronic
10
3,4-trans-Diol derivatives were next synthesized via a 3,4-epoxy
derivative. To our delight, the direction of epoxide formation could
Lipase PS-C
Phosphate Buffer
acetone
be controlled by choosing reaction conditions: the reaction of
with vanadyl acetylacetonate and tert-butyl hydroperoxide gave
3,4-syn-epoxide -20 exclusively, and treatment of -4 with 3-
methyl-3-(trifluoromethyl)dioxirane, generated in situ from triflu-
oroacetone and OxoneÒ, gave 3,4-trans-epoxide
-21 dominantly. It
L-4
40 oC, 3 days
L
L
OAc
OH
L
is likely that the former epoxidation proceeds through a chelation-
controlled mechanism. Deprotection and hydrolytic opening of the
oxirane ring could be done in one step by acid treatment. Thus, 3,4-
N
CO2Et
N
CO2Et
Boc
-10
Boc
L
-5
D
30 %
99 %ee
<57 %
99 %ee
syn-epoxide
mixture of -22 and
could be separated by repeated recrystallization. The dominant
L
-20 was treated with refluxing 5 N H2SO4 to give a 9:1
L
L
-2315 in 92% yield. Fortunately, these products
Scheme 4.

8276
Y. Yoshimura et al. / Bioorg. Med. Chem. 16 (2008) 8273–8286
O
L
-4
OH
OH
H2SO4
H2O
OH
HO
K2OsO4 · 2H2O
NMO
acetone : H2O
r.t., 60 h
100 oC, 5 h
quant.
N
CO2Et
Boc
-20
N
H
COOH
L
L-22
VO(acac)2
(10 mol%)
TBHP
CH2Cl2
r.t., 10 h
92 %
OH
OH
OH
HO
HO
OH
OH
+
OH
HO
N
CO2Et
N
CO2Et
Boc
Boc
N
H
COOH
L
-4
L-16
L
-17
32 %
59 %
CF3COCH3
Oxone®
L
-23
(L-22 : L-23 = 9 : 1)
5N-HCl
NaHCO3
aq.Na2EDTA
CH3CN
5N-HCl
120 oC, 2.5 h
99 %
120 oC, 2.5 h
quant.
0 oC, 30 min
93 %
OH
H2SO4
H2O
OH
O
OH
OH
HO
HO
OH
L
-23
L
-22
100 oC, 4 h
88 %
N
CO2Et
(
L-22 :
L-23= 1 : 4)
N
H
COOH
N
H
COOH
Boc
L
-21
L-18
L-19
Scheme 9.
Scheme 8.
formation of L-22 can simply be explained by the fact that nucleo-
philic attack of water is likely to occur at the less-hindered C-5 po-
sition via trans-diaxial-opening of the epoxide ring. A similar
OH
N
K2OsO4 · 2H2O
NMO
OH
HO
L
-5
reaction of 3,4-trans-epoxide
mixture of -22 and -23, in 88% yield, which are equal to 5-aza-
-altro- and 5-aza- -glucopyranuronic acids, respectively. The for-
-23 was a natural product isolated from the seeds of Baphia
-23 were identical to
those reported. The result suggests that hydrolytic cleavage of
the oxirane ring of -21 also proceeded by way of nucleophilic at-
tack at the less-hindered C-5 position. It is notable that either of
the 5-azasugar derivatives corresponding to -altro- and -glu-
L-21, on the other hand, gave a 1:4
acetone : H2O = 10 : 1
r.t., 23 h
94 %
CO2Et
Boc
L
L
D
D
L-24
mer
L
racemosa.15 All of the instrumental data for
L
5N-HCl
120 oC, 4 h
80 %
L
OH
D
D
OH
HO
copyranuronic acids could selectively be synthesized from the
same intermediate (Scheme 9).
As described in the synthesis of 3-hydroxypipecolic acids, it was
expected that the above-mentioned syntheses could be applied to
N
H
COOH
L-25
the syn-intermediate L-5 to prepare the other four stereoisomers of
5-azapyranuronic acids.
Scheme 10.
An attempt to synthesize 5-aza derivatives of
D
-gulo- and
-5 with potassium
-24, which was further sub-
D-tal-
lopyranuronic acids was made. The reaction of
osmate and NMO gave a sole product
L
tempted to obtain a 3,4-cis-epoxide derivative by the reaction
using vanadyl acetylacetonate. However, the reaction gave a trou-
blesome complex mixture. For the same reason as that described
above, it is thought that the pseudoequatorial hydroxyl group at
the 3-position could no longer be an anchor for the formation of
a vanadyl complex. In contrast, it has been reported that epoxida-
tion using mCPBA more selectively gave a cis-epoxide in a cyclo-
hex-2-enol system when an allylic hydroxyl group occupied a
L
jected to acid hydrolysis. The resulting product was revealed by
instrumental analysis data to be identical to those of the known
D
-isomer of trihydroxypipecolic acid16 and was determined as
L-
25, corresponding to 5-aza-
Many efforts were made to obtain 5-azapyranuronic acid having
-tallo configurations; however, the formation of any trace of the
desired product was not observed. The exclusive formation of
D-gulopyranuronic acid (Scheme 10).
D
L
-
pseudoequatorial rather than
encouraged us to try the reaction of
a 1:7 mixture of -26 and -27 in 91% yield (condition A, Scheme
11). The major product of -5 obtained by trifluoromethyldioxira-
ne-mediated epoxidation was -26, as expected, with a ratio of
6:1 to -26/ -27 (condition B, Scheme 11).
Deprotection and hydrolytic cleavage of epoxide of
treatment of refluxing 5 N HCl dominantly gave 5-aza-
ranuronic acid -28 with a trace amount of -29 in 85% yield. The
a
pseudoaxial position.17 This
-5 with mCPBA, which gave
24 was ascribed to an 1,3-allylic strain between ethoxycarbonyl
and Boc groups which forces the ethoxycarbonyl group to occupy
a pseudoaxial position. As a result, the approaching of the reagent
L
L
L
L
from a b-face of
L-5 was impeded. Owing to the 1,3-allylic strain in
L
L-5 mentioned above, a hydroxyl group at C3 was also forced to oc-
L
L
cupy a pseudoequatorial position, suppressing the process of cis-
dihydroxylation through a chelation-controlled mechanism.
L
-26 by the
-idopy-
D
Finally, the synthesis of 5-aza-
D-ido- and 5-aza-D-galactopy-
L
L
ranuronic acids was tried via epoxy derivatives. First, we at-

Y. Yoshimura et al. / Bioorg. Med. Chem. 16 (2008) 8273–8286
8277
We have also synthesized an
-18, -19, -22, -23, -25,
Section 5).
L
-series of 5-azapyranuronic acids
O
O
OH
OH
D
D
D
D
D
D-28, and D-29 from D-4 and D-5 (see
Conditions
L-5
N
CO2Et
N
CO2Et
Boc
Boc
-26
3. Biological
L
-27
L
Since 3-hydroxypipecolic acid derivatives are designed as aza-
sugar analogues of uronic acid derivatives, they are expected to
act as transition state analogues and to be inhibitors for b-glucu-
ronidase as mentioned earlier. In addition, there are a few reports
on the inhibitory effect of pipecolic acid derivatives on glycosi-
dases. The effect arising by replacement of 5-hydroxymethyl to a
carboxyl group on glycosidase inhibitory activities is also of
interest to us. Thus, we examined inhibitory activities of the ob-
tained 3-hydroxy- and 3,4,5-trihydroxypipecolic acid derivatives
against various glycosidases, including b-glucuronidase. None of
the compounds tested showed inhibitory activity against glucosi-
dases, galactosidases, and mannosidases (data not shown). The re-
sults clearly showed that these enzymes strictly recognize the 5-
hydroxymethyl group. Some of the compounds tested, on one
hand, showed inhibitory activities against b-glucuronidase, b-N-
L
-26 :
L
-27 = 1 :7
Condition A mCPBA, CH2Cl2
r.t., 41 h, 91 %
L
-26 :
L-27 = 6 : 1
Condition B CF3COCH3
Oxone / NaHCO3
aq.Na2EDTA, CH3CN
0 oC, 30 min, 97 %
Scheme 11.
L
-26
aq.H2SO4
84 %
-28 : L-29 = 30 : 1
100 oC, 12 h
L
acetylglucosaminidase, and
sults are summarized in Tables 2 and 3. 3-Hydroxypipecolic acids
-9 and -13 showed moderate inhibitory activity against Esche-
richia coli b-glucuronidase, and their enantiomers -9 and -13 also
showed similar results. Only weak inhibition by -9 and -13
a-N-acetylgalactosaminidase. The re-
L
L
OH
OH
OH
D
D
OH
HO
HO
D
D
against the same enzyme isolated from bovine liver was observed.
Unexpectedly, the 3-hydroxypipecolic acid derivatives showed
weak to moderate inhibitory activity against b-N-acetylglucosa-
N
COOH
N
H
COOH
H
L
-29
L
-28
minidase. It is noteworthy that
L
-7 showed the most potent inhib-
M against the
itory activity: its IC50 values were 720 and 750
l
enzyme isolated from bovine kidney and human placenta,
respectively.
1) TFA,THF-H2O
80 oC, 2 days
2) 5N-HCl
89 %
-28 :
L
L
-29 = 1 : 6
Recently, the dynamic modification of cytosolic and nuclear
proteins by O-glycosylation with b-N-acetylglucosamine (O-Glc-
NAc) on serine and threonine residues has been found to be one
of the post-translational modifications.18 It has also been shown
that O-GlcNAc is stimulated by high glucose flux and implicated
to type II diabetes.19 The cycle of addition/removal of b-N-acetyl-
glucosamine is rapid and is catalyzed by different enzymes similar
to protein phosphorylation/dephosphorylation catalyzed by ki-
nases and phosphatases.20 The enzyme for the addition process
of N-acetylglucosamine is O-GlcNAc transferase, and the removal
process is catalyzed by O-GlcNAcase. The O-GlcNAcase inhibitor
PUGNAc has been used for investigating the biological function
of O-GlcNAc.21 b-N-Acetylglucosaminidase, which is also known
as b-N-acetylhexosaminidase, is functionally different to O-GlcNA-
case.22 However, PUGNAc inhibits both O-GlcNAcase and b-N-acet-
120 oC, 6 h
L
-27
Scheme 12.
result can be interpreted by the fact that the trans-diaxial-opening
of epoxide favoring nucleophilic attack at the C5 position of
gave rise to -28. For a similar reason, the acid treatment of
using the same condition gave a 2:1 mixture of -28/
not shown). Interestingly, when the reaction was performed using
trifluoroacetic acid instead of 5 N H2SO4, 5-aza- -galactopyranu-
ronic acid -29 was dominantly obtained from -5 in 89% yield (
28/ -29 = 1:6, Scheme 12). The reason for the change in selectivity
L
-26
-27
L
L
L
L-29 (data
D
L
L
L-
ylglucosaminidase.23 Moreover, the study to find
inhibitor for O-GlcNAcase is proceeding.24 These facts suggest that
a
selective
L
of the products when a different acid was used is not clear.
Table 2
Inhibition rates of 3-hydroxypipecolic acid derivatives against b-glucuronidase, b-N-acetylglucosaminidase, and
Compound Inhibition rate (IC50 value)
b-N-Acetylglucosaminidase
Bovine kidney Human placenta
a-N-acetylgalactosaminidase at 1000 lM
b-Glucuronidase
Bovine liver
a-N-Acetylgalactosaminidase
E. coli
Chicken liver
L
L
L
L
-7
-9
-11
-13
19%
0.89%
8.6%
2%
58% (720
38%
7.3%
l
M)
60% (750
36%
17%
l
M)
25%
5.2%
0.2%
2.5%
31%
3.3%
24%
0.18%
20%
30%
D
D
D
D
-7
-9
-11
-13
0.2%
0.62%
10%
0.9%
20%
0.9%
23%
15%
32%
0.8%
47%
14%
25%
14%
46%
3.2%
1.3%
1.3%
3.2%
15%

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Y. Yoshimura et al. / Bioorg. Med. Chem. 16 (2008) 8273–8286
Table 3
Inhibition rates of 3,4,5-trihydroxypipecolic acid derivatives against b-glucuronidase, b-N-acetylglucosaminidase, and a-N-acetylgalactosaminidase at 1000 lM
Compound
Configuration
Inhibition rate (IC50 value)
b-N-Acetylglucosaminidase
Bovine kidney Human placenta
b-Glucuronidase
Bovine liver E. coli
a-N-Acetylgalactosaminidase
Chicken liver
L-18
L-19
L-22
L-23
L-25
L-28
L-29
D
D
D
D
D
D
D
-Allo
14%
2.1%
20%
95% (70
8.8%
1%
0.5%
14%
17%
13%
5.5%
4.8%
1.9%
28%
26%
42%
43%
7.6%
6.3%
2.6%
3.8%
4.5%
11%
0.2%
21%
4.2%
31%
-Manno
-Altro
-Gluco
-Gulo
-Ido
-Galacto
0.6%
1.1%
72% (215
2.3%
2.6%
21%
l
l
M)
M)
lM)
17.5%
87% (86
D-18
D-19
D-22
D-23
D-25
D-28
D
-29
L
L
L
L
L
L
L
-Allo
12%
2.6%
1.3%
5.2%
5.4%
1.2%
4.2%
6.3%
18%
10%
27%
11%
1.7%
4.4%
0.5%
39%
26%
42%
50%
2.1%
5.4%
0.8%
4.4%
8.2%
4%
1.3%
7.2%
3.4%
10%
-Manno
-Altro
-Gluco
-Gulo
-Ido
-Galacto
3.1%
6.8%
2.7%
9.4%
15.8%
7.1%
a compound possessing inhibitory activity against b-N-acetylglu-
cosaminidase may also have inhibitory activity against O-GlcNA-
4,5-dehydro-3-hydroxypipecolic acid
activity against the same enzyme. However, the reason for this is
unclear (Table 2).
L
-7 showed a weak inhibitory
case. Therefore, the finding that
against b-N-acetylglucosaminidase is interesting because
be a new lead for designing novel O-GlcNAcase inhibitors.
Stereochemistries of -7 at 2- and 3-positions accord well with
N-acetylglucosamine, and this may lead to a potent inhibitory
activity. In contrast to 4,5-dehydro-3-hydroxypipecolic acids -7
and -11, all stereoisomers of 3-hydroxypipecolic acids -9, -13,
-9, and -13 showed inhibitory activity against b-N-acetylglucosa-
L-7 showed inhibitory activity
L-7 may
In the case of 3,4,5-trihydroxypipecolic acids,
have the most potent inhibitory activity against b-glucuronidase
among the compounds tested. As mentioned above, -23 is a natu-
ral product and was already reported to have an inhibitory activity
against human liver b-glucuronidase.4a The IC50 value of
-23 for E.
coli b-glucuronidase is 215 M, which is comparable to that previ-
ously reported (Table 3).4a For the same enzyme from bovine liver,
-23 showed a more potent inhibitory activity (IC50 = 70 M). The
structure of -23 was designed as a 5-aza analogue of -glucuronic
acid, which should have the function of a transition-state inhibitor
L-23 was found to
L
L
L
L
L
L
L
l
D
D
minidase as well as E. coli b-glucuronidase. Although most of the
compounds tested showed negligible inhibitory activity against
L
l
L
D
a-N-acetylgalactosaminidase obtained from chicken liver, only
Figure 1. Lineweaver–Burk plots of
were 0 (j), 100 M (N). The calculated Ki values were 6.5
M (ꢁ), and 250
calculated Ki values were 400 M.
L
-23 and
L
-29 inhibition of bovine liver b-glucuronidase (a and b) and E. coli b-glucuronidase (c). Concentrations of
L
-23 (a) and
L-29 (b)
l
l
l
M and 15.4 M, respectively. Concentrations of -23 were 0 (j), 250 l
l
L
l
M (ꢁ), and 500
M (N). The
l

Y. Yoshimura et al. / Bioorg. Med. Chem. 16 (2008) 8273–8286
8279
for enzymes recognizing
23 had inhibitory activity against b-glucuronidase meets our
expectation. On the other hand, it is interesting that -29, which
has -galacto configurations as the azasugar, showed an inhibitory
activity against bovine liver b-glucuronidase (IC50 = 86 M) com-
parable to that of -23.
To investigate the mode of inhibition against b-glucuronidase
-23 and -29, the reaction was analyzed by Lineweaver–Burk
plots. From the results shown in Figure 1, it is clear that both
23 and -29 inhibit b-glucuronidase in a competitive manner. The
Ki values of -23 and -29 to the enzyme from bovine liver were
6.5 and 15.4 M, respectively. In addition, the Ki value of -23 to
the same enzyme from E. coli was much weaker (400 M). The fact
that -23 competitively inhibits b-glucuronidases from bovine liver
as well as from E. coli may also be evidence for its action as a tran-
sition-state analogue of -glucuronic acid as mentioned above.
Moreover, the result reveals that -29, similar to -23, can bind to
D
-glucuronic acid. Thus, the finding that
L
-
and allylglycine derivatives, (2) RCM reaction, (3) lipase-catalyzed
kinetic resolution, and (4) additional cis- or trans-dihydroxylation
of alkene. The use of lipase reaction makes it convenient to prepare
both L- and D-enantiomers of pipecolic acid derivatives. As a result,
we have synthesized all stereoisomers of 3-hydroxypipecolic acids,
including 4,5-dehydro-3-hydroxypipecolic acids. Also, the synthesis
L
D
l
L
of L- and D-enantiomers of 3,4,5-trihydroxypipecolic acids having
by
L
L
allo, manno, altro, gluco, gulo, ido, and galacto configurations as aza-
sugars. Among the stereoisomers of 3,4,5-trihydroxypipecolic acids,
only the derivative possessing a tallo configuration could not be ob-
tained. The compounds obtained were evaluated for their inhibitory
activities against various glycosidases, including b-glucuronidase.
None of the compounds tested showed inhibitory activity against
L
-
L
L
L
l
L
l
L
a
- and b-glucosidases.
acid, was found to have potent inhibitory activity against b-glucu-
ronidase as expected. -29, 5-aza-galactopyranuronic acid, also
showed inhibitory activity against b-glucuronidase, comparable to
that of -23. In addition, it is interesting that some uronic-type aza-
L-23, an azasugar analogue of D-glucuronic
D
L
L
L
an active site of b-glucuronidase and act as a transition-state inhib-
itor. The results also suggest that b-glucuronidase should not
strictly recognize C-4 configurations of these inhibitors.
L
sugar derivatives showed moderate inhibitory activities against b-
N-acetylglucosaminidase. These analogues are expected to be new
lead compounds for designing inhibitors for b-glucuronidase as well
as b-N-acetylglucosaminidase.
Inhibitors of b-glucuronidase are expected to become drugs that
suppress side-effects of the camptotecin derivative CPT-11,3 an anti-
tumor drug used for the treatment of small cell lung carcinoma and
other solid tumors. One of the serious side-effects of this drug is diar-
rhea. There are two types of diarrhea induced by CPT-11. Acute diar-
rhea occurring at the early stage of administration is caused by the
anticholinesterase effect of CPT-11.25 Delayed diarrhea occurring
at a later stage is more serious. CPT-11 is changed to an active form
SN-38 by the action of carboxyesterase in the liver. SN-38 is further
changed to inactive SN-38 glucuronate conjugate, which is excreted
into bile.26 By the action of b-glucuronidase from intestinal bacteria,
deconjugation of SN-38 glucuronide gives rise to SN-38, which
causes the severe delayed diarrhea in patients treated with CPT-
11.26 In Japan, Kampo medicine containing baicalin, a b-glucuroni-
dase inhibitor, is co-administrated with CPT-11 to suppress delayed
5. Experimental
Melting points are uncorrected. NMR spectra were recorded at
400 MHz (1H), 100 MHz (13C) using CDCl3, CD3OD, and D2O. As
an internal standard, tetramethylsilane was used for CDCl3 and
CD3OD and 1,4-dioxane was used for D2O. Mass spectra were ob-
tained by EI or FAB mode. Silica gel for chromatography was Fuji
Silysia PSQ 100B. When the reagents sensitive to moisture were
used, the reaction was performed under argon atmosphere.
5.1. tert-Butyl 1-(ethoxycarbonyl)-2-hydroxybut-3-
enylallylcarbamate (3)
diarrhea.3 Therefore, compounds
L-23 and L-29, showing inhibitory
To a solution of 1 M LiHMDS (25 mL, 25 mmol) in THF (200 mL)
was slowly added a solution of 29 (5.0 g, 21 mmol) in THF (100 mL)
at ꢀ80 °C. After being stirred for 30 min, acrolein (2.1 mL, 31 mmol)
was added and the mixture was stirred at the same temperature for
1 h. The reaction mixture was quenched by the addition of satd
NH4Cl and partitioned between AcOEt and H2O. The organic layer
waswashedwithbrineanddriedoverNa2SO4. Afterfiltration, thefil-
trate was concentrated under reduced pressure. The residue was
purified by silica gel column chromatography (n-hexane/ethyl ace-
tate = 9:1) to give 3 (6.4 g, 87%) as an oil: 1H NMR (400 MHz, CDCl3)
d 1.25–1.33 (3H, m), 1.46 (9H, d, J = 10.7 Hz), 3.54–3.70 (1H, m),
3.76–4.03 (1H, m), 4.10–4.28 (3H, m), 4.76–4.82 (1H, m), 5.12–
5.27 (3H, m), 5.33–5.42 (1H, m), 5.77–5.89 (2H, m); 13C NMR
(100 MHz, CDCl3) d 13.7, 13.8, 27.9, 27.9, 51.7, 51.8, 60.9, 61.2,
63.2, 63.8, 71.4, 80.2, 80.6, 80.7, 116.6, 116.8, 117.2, 117.8, 133.6,
133.9, 135.7, 136.5, 137.2, 153.7, 154.8, 171.3, 171.9; IR (neat):
3474.1, 2980.5, 1739.9, 1699.0 cmꢀ1; EI-MS (m/z): 300 (M++1); Anal.
Calcd for C15H25NO5: C, 60.18; H, 8.42; N, 4.68. Found: C, 59.94; H,
8.36; N, 4.71.
activity against b-glucuronidase, are noteworthy. Although they
showed slightly weaker inhibitory activity against the E. coli b-glu-
curonidase than the bovine liver one, they have a possibility of
becoming lead compounds for designing novel therapeutics for
CPT-11-induced diarrhea. From our previous results for azasugars
(DNJ series), some of the
non-competitive inhibitors for glycosidases.2 However, none of the
-enantiomers of 3,4,5-trihydroxypipecolic acid derivatives showed
L-enantiomers of azasugar could act as
L
inhibitory activity against b-glucuronidase.
Against b-N-acetylglucosaminidase, both
L- and D-enantiomers
of 18, 19, 22, and 23, which have 2,3-trans stereochemistry,
showed moderate inhibitory activity. These results are in contrast
to those for 3-hydroxypipecolic acid derivatives: both 2,3-cis and
2,3-trans derivatives have moderate inhibitory activity.
the only derivative that showed a moderate inhibitory activity
against b-N-acetylgalactosaminidase. Indeed, -29 has -galacto
L-29 was
L
D
configuration, but lacks 2-acetamido and 5-hydroxymethyl groups
compared with N-acetylgalactosamine. The inadequate structural
similarity of L-29 to parental N-acetylgalactosamine affects the
inhibitory activity.
5.2. tert-Butyl 2-ethyl 2,3-trans-2,3-dihydro-3-hydroxypyridine-
1,2(6H)-dicarboxylate (4) and tert-butyl 2-ethyl2,3-cis-2,3-
dihydro-3-hydroxypyridine-1,2(6H)-dicarboxylate (5)
4. Conclusion
Our plan was initially made for the synthesis of all stereoisomers
of 3-hydroxy- and 3,4,5-trihydroxypipecolic acid derivatives to sup-
ply samples for SAR study of glycosidase inhibitors. To achieve this,
we have developed a simple, facile, and inexpensive method for
accessing hydroxypipecolic acid derivatives. The method we devel-
oped consists of (1) cross aldol-type condensation between acrolein
To a solution of 3 (6.0 g, 20 mmol) in dry CH2Cl2 (500 mL) was
added Grubbs’ catalyst 1st (166 mg, 0.2 mmol). After the mixture
was stirred at room temperature for 12 h, the solvent was removed
under reduced pressure and the residue was purified by silica gel
column chromatography (n-hexane/ethyl acetate = 4:1) to give
more polar 4 (4.4 g, 80%) and less polar 5 (1.0 g, 19%) as a syrup.

8280
Y. Yoshimura et al. / Bioorg. Med. Chem. 16 (2008) 8273–8286
4: 1H NMR (400 MHz, CDCl3) d 1.26 (3H, t, J = 7.3 Hz), 1.49 (9H,
of L-4 was determined as 99%ee: column: CHIRALPAK IA, hexane/
d, J = 11.6 Hz), 1.87–1.96 (1H, m), 3.78–3.92 (1H, m), 4.09–4.26
(3H, m), 4.60 (0.5H, s), 4.67 (0.5H, s), 4.93 (0.5H, s), 5.06 (0.5H,
s), 5.85–5.99 (2H, m); 13C NMR (100 MHz, CDCl3) d 13.8, 13.8,
27.9, 28.0, 41.5, 42.3, 58.9, 60.3, 60.9, 63.6, 63.8, 80.3, 80.4, 123.8,
123.9, 128.0, 128.2, 155.7, 156.0, 169.1; IR (neat): 3444.6, 2979.2,
1738.9, 1732.4, 1694.6, 1682.7 cmꢀ1; EI-MS (m/z): 271 (M+); HRMS
Calcd for C13H21NO5: 271.1420. Found: 271.1419.
THF = 85:15, flow rate 1.0 mL/min, retention time 26.2 (major) and
31.8 (minor) min at 33 °C, detection: 254 nm.
By the same method, enatiomeric excess of
D-6, after deacetyla-
tion (vide infra), was determined as 97%ee.
(2R,3R)-Ethyl 3-hydroxy-1-tosyl-1,2,3,6-tetrahydropyridine-2-
carboxylate: ½a D28
ꢂ
ꢀ62.7° (c 1.19, CHCl3).
5: 1H NMR (400 MHz, CDCl3) d 1.27 (3H, t, J = 7.3 Hz), 1.49 (9H,
d, J = 1.9 Hz), 3.54–3.66 (1H, m), 3.97–4.27 (4H, m), 4.44–4.46 (1H,
m), 5.04 (0.5H, d, J = 5.3 Hz), 5.21 (0.5H, d, J = 5.3 Hz), 5.61–5.69
(1H, m), 5.88 (1H, d, J = 10.6 Hz); 13C NMR (100 MHz, CDCl3) d
13.6, 13.7, 27.8, 41.4, 42.1, 54.1, 55.6, 61.0, 65.6, 80.2, 80.3,
123.2, 123.4, 129.3, 129.7, 154.1, 154.6, 171.2, 171.3; IR (neat):
3475.0, 2979.2, 1739.7, 1704.7 cmꢀ1; EI-MS (m/z): 271 (M+); HRMS
Calcd for C13H21NO5: 271.1420. Found: 271.1432.
5.5. (2S,3S)-3-Hydroxybaikiain (L-7)
A mixture of L-4 (300 mg, 1.11 mmol) and 5 N HCl (32 mL) was
kept at 120 °C for 3 h. The solvent was removed under reduced
pressure and the residue was dissolved in H2O (50 mL). The mix-
ture was applied to a top of ion-exchange column (DOWEX 50W
X8) and eluted with H2O and then with 0.5 N NH3. The eluate of
0.5 N NH3 was concentrated to dryness under reduced pressure
to give
½ ꢂ
L
-7 (157 mg, 99%) as a crystal: mp 232–237 °C (decomp.);
5.3. (2R,3R)-1-tert-Butyl 2-ethyl 3-acetoxy-2,3-dihydropyridine-
a 2D6
+60.6° (c 1.0, H2O); 1H NMR (400 MHz, D2O) d 3.60 (1H, dd,
1,2(6H)-dicarboxylate (D-6) and (2S,3S)-1-tert-butyl 2-ethyl 3-
J = 17.4, 2.4 Hz), 3.68 (1H, d, J = 5.8 Hz), 3.74 (1H, dd, J = 17.4,
2.4 Hz), 4.48–4.51 (1H, m), 5.77–5.81 (1H, m), 5.85–5.89 (1H, m);
13C NMR (100 MHz, D2O) d 40.7, 60.3, 63.1, 123.2, 127.1, 171.8;
IR (KBr): 3403, 1647, 1409, 1091, 1059 cmꢀ1; EI-MS (m/z): 143
(M+); Anal. Calcd for C6H9NO3ꢃ0.15H2O: C, 49.41; H, 6.43; N, 9.60.
Found: C, 49.44; H, 6.36; N, 9.64.
hydroxy-2,3-dihydropyridine-1,2(6H)-dicarboxylate (
L-4)
To a solution of 4 (2.1 g, 7.6 mmol) in diisopropyl ether (33 mL)
were added lipase PS-C Amano II (3.8 g, purchased from Wako Co.
Ltd) and vinyl acetate (65 mL, 708 mmol). The mixture was kept at
40 °C for 3 days. Insoluble materials were removed by suction and
the filtrate was concentrated under reduced pressure. The residue
was purified by silica gel column chromatography (n-hexane/ethyl
5.6. (2S,3S)-1-tert-Butyl 2-ethyl 3-hydroxypiperidine-1,2-dicar-
boxylate (L-8)
acetate = 4:1) to give less polar D-6 (1.1g, 47%, and 97%ee) and
more polar
L
-4 (978 mg, 47%, 99%ee).
A mixture of L-4 (518 mg, 1.9 mmol) and Pd–C (5%) (129 mg) in
D
-6: syrup; ½a 2D1
ꢂ
ꢀ163.3° (c 1.0, CHCl3); 1H NMR (400 MHz,
AcOEt (52 mL) was stirred at room temperature for 15 h under H2
atmosphere. Insoluble materials were removed by Celite filtration
and the filtrate was concentrated under reduced pressure. The res-
idue was purified by silica gel column chromatography (n-hexane/
CDCl3) d 1.24–1.28 (3H, m), 1.49 (9H, d, J = 16.9 Hz), 2.07 (3H, s),
3.80 (0.5H, d, J = 17.9, 1.9 Hz), 3.90 (0.5H, d, J = 19.3, 1.9 Hz),
4.15–4.26 (3H, m), 5.04 (0.5H, d, J = 1.9 Hz), 5.15 (0.5H, d,
J = 1.9H), 5.61 (0.5H, d, J = 5.8 Hz), 5.68 (0.5H, d, J = 5.3 Hz), 5.93–
6.07 (2H, m); 13C NMR (100 MHz, CDCl3) d 14.0, 14.0, 20.9, 20.9,
28.2, 28.2, 41.3, 42.2, 55.9, 57.0, 61.5, 66.0, 66.2, 80.5, 80.6, 120.2,
120.5, 130.1, 130.9, 131.4, 154.8, 155.7, 168.3, 168.4, 170.2; IR
(neat): 2979.5, 1732.1, 1694.9, 1668.0 cmꢀ1; EI-MS (m/z): 313
(M+); HRMS Calcd for C15H23NO6: 313.1525. Found: 313.1523.
ethyl acetate = 4:1) to give L-8 (514 mg, 99%) as a crystal: mp 86–
88 °C; ½a 2D1
ꢂ
ꢀ31.5° (c 1.00, CHCl3); 1H NMR (400 MHz, CDCl3) d 1.28
(3H, t, J = 7.0 Hz), 1.46–1.53 (11H, m), 1.79–1.82 (2H, m), 2.26
(0.5H, br s), 2.38 (0.5H, br s), 2.91 (0.5H, br s), 3.05 (0.5H, br s),
3.91 (0.5H, br s), 4.01 (0.5H, br s), 4.21 (2H, q, J = 7.0 Hz), 4.42
(1H, s), 4.73 (0.5H, br s), 4.79 (0.5H, br s); 13C NMR (100 MHz,
CDCl3) d 14.0, 18.0, 27.3, 28.1, 40.6, 41.7, 60.2, 60.9, 65.0, 80.0,
156.0, 156.4, 169.7; IR (neat): 3445.1, 2978.5, 1739.2,
1682.9 cmꢀ1; EI-MS (m/z): 273 (M+); Anal. Calcd for C13H23NO5:
C, 57.13; H, 8.48; N, 5.12. Found: C, 57.14; H, 8.57; N, 5.08.
L
-4: white crystal; mp 61–63 °C; ½a D21
ꢂ
+52.5° (c 0.99, CHCl3).
-4 and -6
-4 (60 mg, 0.22 mmol) in CH2Cl2 (5 mL) was
5.4. Determination of optical purity of
L
D
To a solution of
L
added TFA (0.8 mL, 10.5 mmol). After the mixture was stirred at
room temperaturefor 1 h, the solvents were removed under reduced
pressure. The residue was dissolved in 1,4-dioxane/H2O = 1:1
(5 mL). To this solution were added p-toluenesulfonyl chloride
(70 mg, 0.22 mmol) and NaHCO3 (278 mg, 3.3 mmol). The mixture
was stirred at room temperature for 3 h diluted with AcOEt, and
washed with H2O. The water layer was extracted by AcOEt three
times and the combined organic layer was washed with brine and
then dried (Na2SO4). After filtration, the filtrate was concentrated
under reduced pressure. The residue was purified by silica gel col-
umn chromatography (n-hexane/ethyl acetate = 4:1) to give
(2S,3S)-ethyl 3-hydroxy-1-tosyl-1,2,3,6-tetrahydropyridine-2-car-
5.7. (2S,3S)-3-Hydroxypipecolic acid (L-9)
By the same procedure for the synthesis of
L
-7,
-8 (249 mg, 0.91 mmol). Mp 244–251 °C
+12.8° (c 0.85, H2O) [lit.686%ee: [
+11.6° (c
L-9 (111 mg,
84%) was obtained from
L
(decomp.); ½a 2D1
ꢂ
a]
D
0.8, H2O)]; 1H NMR (400 MHz, D2O) d 1.47–1.60 (2H, m), 1.74–
1.89 (2H, m), 2.90–2.97 (1H, m), 3.16–3.22 (1H, m), 3.45 (1H, d,
J = 6.8 Hz), 3.97–4.01 (1H, m); 13C NMR (100 MHz, D2O) d 18.3,
28.1, 42.5, 61.7, 65.8, 171.6; IR (KBr): 3282.8, 2983.9, 2959.4,
1624.1, 1599.1, 1401.3 cmꢀ1; FAB-MS (m/z): 146 (M++1); Anal.
Calcd for C8H11NO3ꢃ1.1H2O: C, 43.68; H, 8.06; N, 8.49. Found: C,
43.66; H, 7.88; N, 8.47.
boxylate (59 mg, 82%). ½a D25
ꢂ
+64.8° (c 1.10, CHCl3); 1H NMR
(400 MHz, CDCl3) d 1.09 (3H, t, J = 7.2 Hz), 2.26 (1H, br s), 2.42 (3H,
s), 3.86–3.92 (2H, m), 3.97–4.05 (2H, m), 4.09–4.15 (1H, m), 4.52
(1H, br s), 4.89 (1H, d, J = 1.9 Hz), 5.87–5.91 (1H, m), 5.94–5.98
(1H, m), 7.30 (2H, d, J = 8.2 Hz), 7.73 (2H, d, J = 8.2 Hz); 13C NMR
(100 MHz, CDCl3) d 13.8, 21.5, 42.5, 60.8, 61.4, 64.7, 124.8, 127.4,
128.0, 129.5, 135.8, 143.7, 168.0; IR (neat): 3501.0, 2982.9, 1746.8,
1732.2, 1335.4, 1162.5 cmꢀ1; EI-MS (m/z): 325 (M+); HRMS Calcd
for C15H19NO5S: 325.098. Found: 325.0989. By HPLC analysis of the
compound thus obtained using chiral column, enatiomeric excess
5.8. tert-Butyl 2-ethyl 3-acetoxy-2,3-dihydropyridine-1,2(6H)-
dicarboxylate (10)
To a solution of 4 (4.5 g, 17 mmol) in THF (88 mL) were added
PPh3 (8.7 g, 33 mmol), AcOH (2.4 mL, 41 mmol), and DEAD (15 mL,
33 mmol) at 0 °C. After the mixture was stirred at room temperature
for 4 h, the solvent was removed under reduced pressure. The resi-
due was purified by silica gel column chromatography (n-hexane/
ethyl acetate = 9:1) to give 10 (<4.6 g, <90%) containing small

Y. Yoshimura et al. / Bioorg. Med. Chem. 16 (2008) 8273–8286
8281
amounts of inseparable impurity: 1H NMR (400 MHz, CDCl3) d 1.26
5.12. (2S,3R)-3-Hydroxypipecolic acid (
L-13)
(3H, t, J = 7,3 Hz), 1.47 (9H, s), 2.10 (3H, s), 3.91–4.18 (4H, m), 5.22
(0.5H, d, J = 5.3 Hz), 5.38 (0.5H, d, J = 5.3 Hz), 5.49 (1H, s), 5.64 (1H,
d, J = 10.6 Hz), 5.86–5.93 (1H, m); 13C NMR (100 MHz, CDCl3) d
14.0, 20.7, 28.0, 41.7, 42.3, 51.5, 53.0, 60.7, 66.0, 80.6, 80.8, 121.9,
122.2, 122.3, 126.3, 126.5, 154.2, 154.7, 169.0, 169.5, 169.6, 169.9;
IR (neat): 3452.2, 2981.8, 1747.5, 1713.5, 1370.8, 1232.1, 1163.4,
1030.4, 755.9 cmꢀ1; EI-MS (m/z): 313 (M+); HRMS Calcd for
By the same procedure for the synthesis of
L
-7,
L
-13 (82 mg,
86%) was obtained from L-12 (168 mg, 0.66 mmol) as a crystal.
mp 270–274 °C (decomp.);
½
a 2D7
ꢂ
ꢀ60.0° (c 0.57, H2O) [lit.27
82%ee: [
a]
ꢀ52.8° (c 0.6, H2O)]; 1H NMR (400 MHz, D2O) d 1.55–
D
1.66 (2H, m), 1.77–1.89 (2H, m), 2.81–2.89 (1H, m), 3.28 (1H, dd,
J = 12.7, 2.4 Hz), 3.52 (1H, d, J = 1.4 Hz), 4.35 (1H, s); 13C NMR
(100 MHz, D2O) d 16.7, 29.5, 44.4, 63.1, 64.9, 173.1; IR (KBr):
3400.8, 3168.6, 3053.8, 1623.0, 1408.2, 1393.2 cmꢀ1; FAB-MS (m/
z): 146 (M++1); HRMS Calcd for C6H12NO3: 146.0817. Found:
146.0820; Anal. Calcd for C6H11NO3: C, 49.65; H, 7.64; N, 9.65.
Found: C, 49.30; H, 7.80; N, 9.59.
C15H23NO6: 313.1525. Found: 313.1529.
5.9. (2R,3S)-1-tert-Butyl 2-ethylcetoxy-2,3-dihydropyridine-
1,2(6H)-dicarboxylate ( -10) and (2S,3R)-1-tert-butyl 2-ethyl
2,3-dihydro-3-hydroxypyridine-1,2(6H)-dicarboxylate ( -5)
D
L
To a solution of 10 (206 mg, 0.66 mmol) were added Lipase PS-C
(340 g) and 0.1 mol/L phosphate buffer solution (pH 7) (7.0 mL).
The mixture was kept at 40 °C for 3 days. Insoluble materials were
removed by Celite filtration and the filtrate was extracted with
AcOEt three times. The combined organic layer was washed with
brine and dried over Na2SO4. After filtration, the filtrate was con-
centrated under reduced pressure. The residue was purified by sil-
ica gel column chromatography (n-hexane/ethyl acetate = 4:1) to
5.13. (2R,3R)-1-tert-Butyl 2-ethyl 3-hydroxy-2,3-dihydro-
pyridine-1,2(6H)-dicarboxylate (
D-4)
To a solution of -6 (50 mg, 0.16 mmol) in MeOH (1.2 mL) was
D
added a solution of LiOH (19 mg, 0.80 mmol) in H2O (0.4 mL) at
ꢀ30 °C. After being stirred at the same temperature for 10 min,
the mixture was neutralized by AcOH and then allowed to warm
to room temperature. The whole was extracted with CHCl3 four
times and the combined organic layer was washed with brine
and then dried (Na2SO4). After filtration, the filtrate was concen-
trated under reduced pressure. The residue was purified by silica
gel column chromatography (n-hexane/ethyl acetate = 1:1) to give
give less polar
amounts of inpurity) and
D
-10 (<118 mg, <57%, and 99%ee, containing small
-5 (55 mg, 30%, and 99%ee). The optical
L
purities of the products were determined after converting an N-to-
syl derivative as described above.
L
-5: ½a 2D1
ꢂ
ꢀ30.9° (c 1.20, CHCl3).
D
-4 (42 mg, 97%). ½a D21
ꢀ54.1° (c 0.98, CHCl3).
ꢂ
N-Tosyl derivative [(2S,3R)-Ethyl 3-hydroxy-1-tosyl-1,2,3,6-
tetrahydropyridine-2-carboxylate]: ½a D28
ꢂ
ꢀ3.4° (c 1.53, CHCl3); 1H
5.14. (2R,3S)-1-tert-Butyl 2-ethyl2,3-dihydro-3-hydroxypyridine-
1,2(6H)-dicarboxylate ( -5), tert-butyl ethyl pyridine-1,2(6H)-
dicarboxylate (14) and (2R,3S)-1-tert-butyl 2-methyl 2,3-dihydro-
3-hydroxypyridine-1,2(6H)-dicarboxylate ( -15)
NMR (400 MHz, CDCl3) d 1.04 (3H, t, J = 7.3 Hz), 2.43 (3H, s), 3.96
(1H, dd, J = 17.4, 2.4 Hz), 3.85–3.93 (1H, m), 3.97–4.08 (2H, m),
4.48 (1H, s), 4.96 (1H, d, J = 5.8 Hz), 5.67 (1H, dd, J = 10.1, 2.4 Hz),
5.80 (1H, d, J = 10.1 Hz), 7.30 (2H, d, J = 7.7 Hz), 7.69 (2H, d,
J = 7.7 Hz); 13C NMR (100 MHz, CDCl3) d 1.0, 13.7, 21.5, 29.7,
42.4, 56.0, 61.6, 66.4, 123.3, 127.3, 129.6, 129.8, 136.1, 143.7,
170.2; EI-MS (m/z): 326 (M++1); HRMS Calcd for C15H19NO5S:
325.0984. Found: 325.0981; HPLC: CHIRALPAK IA, hexane/
THF = 85:15, flow rate 1.0 mL/min, retention time 20.5 (minor),
32.5 (major) min, 34 °C, detection: 254 nm.
D
D
To a solution of D-10 (500 mg, 1.6 mmol) in MeOH (12 mL) was
added a solution of LiOH (196 mg, 8.0 mmol) in H2O (4 mL) at
ꢀ30 °C. After being stirred at the same temperature for 30 min, the
mixturewas neutralized by AcOH and then allowedto warmto room
temperature. The whole mixture was extracted with CHCl3 four
times and the combined organic layer was washed with brine and
then dried (Na2SO4). After filtration, the filtrate was concentrated
under reduced pressure. The residue was purified by silica gel col-
5.10. (2S,3R)-3-Hydroxybaikiain (
L-11)
umn chromatography (n-hexane/ethyl acetate = 9:1) to give
D-4
By the same procedure as for the synthesis of
L
-7,
L
-11 (61 mg,
(261 mg, 60%), 14 (48 mg, 12%), and D-15 (8 mg, 2%), respectively.
85%) was obtained from
L
-5 (136 mg, 0.50 mmol) as a crystal. mp
D
-4: ½a 2D1
ꢂ
+29.6° (c 0.94, CHCl3).
+0.69° (c 0.95, CHCl3); 1H NMR (400 MHz, CDCl3) d 1.32
285–287 °C; ½a 2D6
ꢂ
ꢀ335.3° (c 1.0, H2O) [lit.6
½
a 2D0
ꢂ
ꢀ332.7° (c 0.3,
14: ½a 2D7
ꢂ
H2O) ]; 1H NMR (400 MHz, D2O) d 3.54–3.60 (1H, m), 3.64 (1H, dd,
J = 4.3, 1.4 Hz), 3.70 (1H, d, J = 2.9 Hz), 4.48 (1H, t, J = 3.9 Hz), 5.82–
5.86 (1H, m), 5.99–6.04 (1H, m); 13C NMR (100 MHz, D2O) d 43.1,
61.5, 61.6, 124.4, 127.6, 172.3; IR (KBr): 3325.1, 3003.9, 2391.4,
1647.0, 1397.3, 1091.8 cmꢀ1; EI-MS (m/z): 143 (M+); Anal. Calcd
for C6H9NO3ꢃ0.1H2O: C, 49.72; H, 6.40; N, 9.66. Found: C, 49.46; H,
6.40; N, 9.48.
(3H, t, J = 7.3 Hz), 1.45 (9H, s), 4.25 (2H, q, J = 7.3 Hz), 4.29–4.30
(2H, m), 5.90–5.94 (1H, m), 6.05–6.10 (1H, m), 6.44 (1H, d,
J = 4.9 Hz); 13C NMR (100 MHz, CDCl3) d 14.1, 27.9, 42.7, 60.9,
81.6, 120.0, 122.5, 127.2, 130.5, 152.6, 164.1; IR (neat): 2980.7,
1714.9, 1355.0, 1275.0, 1166.9, 1075.4, 764.3 cmꢀ1; EI-MS (m/z):
253 (M+); HRMS Calcd for C13H19NO4: 253.1314. Found: 253.1302.
D
-15: ½a 2D7
ꢂ
ꢀ37.6° (c 0.65, CHCl3); 1H NMR (400 MHz, CDCl3) d
1.49 (9H, s), 3.49–3.67 (1H, m), 3.75 (3H, s), 3.94–4.10 (2H, m),
4.45 (1H, br s), 5.06 (0.5H, d, J = 5.3 Hz), 5.25 (0.5H, d, J = 5.3 Hz),
5.61–5.69 (1H, m), 5.87 (1H, d, J = 10.6 Hz); 13C NMR (100 MHz,
CDCl3) d 28.2, 41.6, 42.3, 52.4, 54.3, 55.9, 66.0, 66.1, 80.8, 123.6,
123.9, 129.8, 130.2, 154.3, 154.9, 172.3; IR (neat): 3480.5, 2977.3,
1743.9, 1695.5, 1404.1, 1164.1, 1005.9, 893.8 cmꢀ1; EI-MS (m/z):
257 (M+); HRMS Calcd for C12H19NO5: 257.1263. Found: 257.1254.
5.11. (2S,3R)-1-tert-Butyl 2-ethyl3-hydroxypiperidine-1,2-dicar-
boxylate (L-12)
By the same procedure as for the synthesis of L-8, -12 (191 mg,
88%) was obtained from
L
-5 (216 mg, 0.80 mmol). ½aLꢂD21 ꢀ60.2° (c
1.22, CHCl3); 1H NMR (400 MHz, CDCl3) d 1.33 (3H, t, J = 7.2 Hz),
1.47 (11H, s), 1.71 (1H, br s), 2.01 (1H, d, J = 11.6 Hz), 2.58–2.74
(1H, m), 3.72 (2H, s), 3.88–4.02 (1H, m), 4.26 (2H, d, J = 7.2 Hz),
4.92–5.06 (1H, m); 13C NMR (100 MHz, CDCl3) d 14.2, 23.5, 23.7,
28.3, 30.3, 40.2, 41.4, 57.2, 58.3, 61.5, 68.9, 69.1, 80.5, 154.8,
154.9, 172.1; IR (neat): 3454.2, 2979.4, 1732.8, 1694.9, 1393.4,
1368.0, 1152.8 cmꢀ1; EI-MS (m/z): 273 (M+); HRMS Calcd for
5.15. (2S,3R,4R,5R)-1-tert-Butyl 2-ethyl 3,4,5-trihydroxypiper-
idine-1,2-dicarboxylate (L-16) and (2S,3R,4S,5S)-1-tert-butyl 2-
ethyl 3,4,5-trihydroxypiperidine-1,2-dicarboxylate (
L
-17)
To a solution of
L-4 (504 mg, 1.9 mmol) in acetone (14 mL) were
C13H23NO5: 273.1576. Found: 273.1562.
added NMO (1.5 mL) and a solution of K2OsO4ꢃ2H2O (6.9 mg,

8282
Y. Yoshimura et al. / Bioorg. Med. Chem. 16 (2008) 8273–8286
0.019 mmol) in H2O (1.4 mL) at 0 °C. The mixture was stirred at
room temperature for 60 h. The reaction was quenched by the
addition of solid Na2S2O3 (871 mg). The mixture was stirred for
30 min and dried over Na2SO4. After filtration, the filtrate was con-
centrated under reduced pressure. The residue was purified by sil-
ica gel column chromatography (CHCl3/MeOH = 19:1) to give more
phy (n-hexane/ethyl acetate = 4:1) to give L-20 (133 mg, 92%) as
a syrup: ½a 2D6
ꢂ
ꢀ10.3° (c 1.17, CHCl3); 1H NMR (400 MHz, CDCl3) d
1.28 (3H, t, J = 6.8), 1.47 (9H, d, J = 8.2 Hz), 2.70 (1H, d,
J = 10.6 Hz), 3.28–3.48 (2H, m), 3.52 (1H, t, J = 4.3 Hz), 4.17 (2H,
q, J = 6.8 Hz), 4.34 (1H, d, J = 15.5 Hz), 4.52 (1H, br s), 4.71 (0.5H,
s), 4.87 (0.5H, s); 13C NMR (100 MHz, CDCl3) d 13.8, 27.8, 38.7,
39.7, 51.1, 51.7, 57.7, 59.0, 61.2, 62.1, 80.8, 155.2, 155.6, 168.8;
IR (neat): 3472.6, 2979.5, 2934.4, 1738.7, 1699.1, 1367.0, 1250.5,
1058.3 cmꢀ1; EI-MS (m/z): 287 (M+); HRMS Calcd for C13H21NO6:
287.1369. Found: 287.1378.
polar
L
-16 (179 mg, 32%) and less polar
L
-17 (336 mg, 59%).
L
-16: ½a 2D7
ꢂ
+6.4° (c 0.93, CHCl3); 1H NMR (400 MHz, CD3OD) d
1.22 (3H, t, J = 6.8 Hz), 1.41 (9H, d, J = 21.3 Hz), 3.06–3.21 (1H,
m), 3.78–3.87 (3H, m), 4.11 (2H, q, J = 6.8 Hz), 4.38 (1H, d,
J = 12.6 Hz), 4.55 (1H, d, J = 24.6 Hz); 13C NMR (100 MHz, CD3OD)
d 14.4, 14.4, 28.5, 28.6, 41.9, 43.0, 58.4, 59.6, 62.1, 65.0, 65.0,
71.2, 71.6, 71.8, 79.4, 81.6, 157.7, 157.8, 170.8, 170.8; IR (neat):
3418.2, 2979.0, 2938.1, 2530.3, 1731.9, 1673.9, 1416.5, 1164.2,
1026.8 cmꢀ1; EI-MS (m/z): 305 (M+); HRMS Calcd for C13H23NO7:
305.1475. Found: 305.1480.
5.19. (1S,2R,3S,6R)-4-tert-Butyl 3-ethyl 2-hydroxy-7-oxa-4-
azabicyclo[4.1.0]heptane-3,4-dicarboxylate (L-21)
To a solution of L-4 (500 mg, 1.8 mmol) in CH3CN (14 mL) were
added 4 ꢄ 10ꢀ4 M Na2EDTA (9.2 mL) and CF3COCH3 (1.8 mL) at
L
-17: ½a 2D7
ꢂ
ꢀ5.9° (c 3.22, CHCl3); 1H NMR (400 MHz, CD3OD) d
0 °C. A mixture of NaHCO3 (1.2 g) and OxoneÒ (5.8 g) was slowly
1.22 (3H, t, J = 6.8 Hz), 1.42 (9H, s), 3.12 (1H, br s), 3.37 (1H, t,
J = 2.9 Hz), 3.83 (1H, br s), 4.12 (1H, s), 4.17 (2H, q, J = 6.8 Hz),
4.27 (1H, br s), 4.94 (1H, br s); 13C NMR (100 MHz, CD3OD) d
14.5, 28.5, 49.9, 55.9, 62.6, 68.1, 70.2, 71.7, 81.9, 158.1, 170.4; IR
(neat): 3390.9, 2979.2, 2934.9, 1739.0, 1698.9, 1417.5, 1368.1,
1250.6, 1091.4 cmꢀ1; EI-MS (m/z): 305 (M+); HRMS Calcd for
added to the mixture of L-4 at 0 °C over 1 h. The mixture was stir-
red at the same temperature for 30 min. H2O was added to the
mixture and the whole mixture was extracted with CH2Cl2 three
times. The combined organic layer was washed with brine and
dried over Na2SO4. After filtration, the filtrate was concentrated
under reduced pressure. The residue was purified by silica gel col-
C
13H23NO7: 305.1475. Found: 305.1480.
umn chromatography (n-hexane/ethyl acetate = 3:2) to give
L
-21
(493 mg, 93%) as a syrup: ½a D27
ꢂ
+19.7° (c 0.99, CHCl3); 1H NMR
5.16. (2S,3R,4R,5R)-3,4,5-Trihydroxypipecolic acid (1-deoxy-5-
aza- -allopyran-uronic acid, -18)
(400 MHz, CDCl3) d 1.24–1.30 (3H, m), 1.46 (9H, d, J = 15.5 Hz),
3.03 (0.5H, d, J = 9.2 Hz), 3.24–3.31 (2.5H, m), 3.78 (1H, dd,
J = 18.8, 15.5 Hz), 3.93 (1H, dt, J = 15.5, 2.9 Hz), 4.16–4.22 (2H,
m), 4.58 (0.5H, d, J = 2.9 Hz), 4.68 (0.5H, d, J = 2.9 Hz), 4.81 (0.5H,
d, J = 9.2 Hz), 4.89 (0.5H, t, J = 3.4 Hz); 13C NMR (100 MHz, CDCl3)
d 14.1, 14.1, 28.2, 28.3, 39.5, 40.1, 49.9, 50.1, 51.4, 51.7, 56.8,
58.2, 61.2, 61.2, 66.1, 66.6, 81.1, 81.2, 155.9, 156.3, 168.4, 168.5;
IR (neat): 3430.2, 2980.2, 2935.9, 1668.4, 1367.7, 1318.1, 1253.2,
1167.9, 1028.1 cmꢀ1; FAB-MS (m/z): 288 (M++1); HRMS Calcd for
D
L
A mixture of L-16 (90 mg, 0.29 mmol) and 5 N HCl (8 mL) was
kept at 120 °C for 2.5 h. The solvent was removed under reduced
pressure and the residue was purified by silica gel column chroma-
tography (MeOH/10% NH3 aq = 30:1) to give L-18 (51 mg, 99%) as a
crystal. Mp 231–234 (decomp.); ½a D27
ꢂ
ꢀ29.7° (c 1.0, H2O) [lit.14
½ ꢂ
a 2D0
ꢀ28.3° (c 0.64, H2O)]; 1H NMR (400 MHz, D2O) d 3.05 (1H, dd,
J = 13.5, 2.4 Hz), 3.26 (1H, dd, J = 13.5, 4.8 Hz), 3.35 (1H, d,
J = 9.2 Hz), 3.60 (1H, dd, J = 9.2, 2.9 Hz), 3.93 (1H, t, J = 9.2 Hz),
4.05–4.07 (1H, m); 13C NMR (100 MHz, D2O) d 46.6, 61.7, 66.2,
68.7, 72.6, 172.7; IR (KBr): 3448.7, 3258.2, 1603.8, 1423.2,
1254.8, 1140.5, 1096.4 cmꢀ1; FAB-MS (m/z): 178 (M++1); Anal.
Calcd for C6H11NO5ꢃ1.2H2O: C, 36.25; H, 6.79; N, 7.05. Found: C,
36.34; H, 6.88; N, 7.01.
C13H22NO6: 288.1447. Found: 288.1455.
5.20. (2S,3R,4S,5R)-3,4,5-Trihydroxypipecolic acid (1-deoxy-5-
aza- -altropyran-uronic acid, -22) and (2S,3R,4R,5S)-3,4,5-
trihydroxypipecolic acid (1-deoxy-5-aza- -glucopyranuronic
acid, -23)
D
L
D
L
From L-20: a mixture of L-20 (340 mg, 1.2 mmol), H2O (4.8 mL)
5.17. (2S,3R,4S,5S)-3,4,5-Trihydroxypipecolic acid (1-deoxy-5-
aza-D-mannopyran-uronic acid, L-19)
and H2SO4 (0.52 mL) was kept at 120 °C for 5 h. The mixture was
allowed to cool to room temperature and was applied to a top of
ion-exchange column (DOWEX 50W X8). The eluate of 0.5 N NH3
was concentrated to dryness under reduced pressure to give a
By the same procedure as for the synthesis of
L
-18,
-17 (55 mg, 0.18 mmol) as a crystal. Mp
+19.2° (c 1.0, H2O); 1H NMR (400 MHz,
L-19 (32 mg,
quant.) was obtained from
L
9:1 mixture of
recrystallization of the 9:1 mixture from MeOH/acetone/H2O gave
analytically pure -22 (77 mg).
From -21: by the same procedure as described above, a 1:4
mixture of -22/ -23 (58 mg, 88%) was obtained from -21
L-22/L-23 (211 mg, quant.). Crystallization and
229–238 °C (decomp.); ½a D26
ꢂ
D2O) d 3.01 (1H, dd, J = 12.6, 9.2 Hz), 3.17 (1H, dd, J = 12.6, 4.3 Hz),
3.60 (1H, d, J = 9.2 Hz), 3.86–3.92 (3H, m); 13C NMR (100 MHz, D2O)
d 43.1, 57.9, 65.9, 69.1, 69.3, 172.8; IR (KBr): 3402.2, 3222.4, 2473.2,
1612.5, 1400.3, 1372.5, 1154.1, 942.3 cmꢀ1; FAB-MS (m/z): 178
(M++1); Anal. Calcd for C6H11NO5ꢃ0.2H2O: C, 39.87; H, 6.36; N, 7.75.
Found: C, 39.69; H, 6.46; N, 7.62.
L
L
L
L
L
(104 mg, 0.36 mmol). Repeated recrystallizations from MeOH/ace-
tone/H2O gave analytically pure -23 (17 mg).
L
-22: mp 232–236 °C; ½a D26
ꢂ
Lꢀ10.3° (c 1.0, H2O); 1H NMR
(400 MHz, D2O) d 3.06 (1H, dd, J = 13.0, 7.7 Hz), 3.26 (1H, dd,
J = 13.0, 3.9 Hz), 3.60 (1H, dd, J = 6.8, 2.9 Hz), 3.80 (1H, d,
J = 6.8 Hz), 3.93–3.98 (1H, m),4.25 (1H, d, J = 6.8, 2.9 Hz); 13C NMR
(100 MHz, D2O) d 44.8, 60.3, 65.7, 68.2, 70.6, 171.6; IR (KBr):
3490.1, 3067.3, 1590.6, 1424.4, 1399.8, 1077.4, 710.8 cmꢀ1; FAB-
MS (m/z): 178 (M++1); Anal. Calcd for C6H11NO5ꢃH2O: C, 36.92; H,
6.71; N, 7.18. Found: C, 36.88; H, 6.70; N, 7.20.
5.18. (1R,2R,3S,6S)-4-tert-Butyl 3-ethyl 2-hydroxy-7-oxa-4-
azabicyclo[4.1.0]heptane-3,4-dicarboxylate (
L-20)
To a solution of -4 (136 mg, 0.50 mmol) in CH2Cl2 (15 mL) were
L
added vanadyl acetylacetonate (14 mg, 0.05 mmol) and t-BuOOH
(0.35 mL) at 0 °C. The mixture was stirred at room temperature
for 10 h. To the mixture, satd Na2S2O3 was added and the whole
was extracted with CH2Cl2 three times. The combined organic layer
was washed with satd NaHCO3 and brine and then dried (Na2SO4).
After filtration, the filtrate was concentrated under reduced pres-
sure. The residue was purified by silica gel column chromatogra-
L
-23: mp 266–272 °C; ½a D26
ꢂ
+19.5° (c 0.79, H2O) [lit.15
[a] +18°
D
(c 0.01, H2O)]; 1H NMR (400 MHz, D2O) d 2.81 (1H, dd, J = 12.6,
11.6 Hz), 3.34–3.42 (3H, m), 3.58 (1H, dd, J = 10.1, 8.7 Hz), 3.64–
3.71 (1H, m); 13C NMR (100 MHz, D2O) d 45.7, 61.6, 67.5, 70.7,
76.3, 172.3; IR (KBr): 3421.6, 1652.0, 1595.1, 1403.6, 1101.7,

Y. Yoshimura et al. / Bioorg. Med. Chem. 16 (2008) 8273–8286
8283
1047.2, 542.5 cmꢀ1; FAB-MS (m/z): 178 (M++1); Anal. Calcd for
L
-27: mp 83–84 °C; ½a D26
ꢂ
ꢀ50.5° (c 0.80, CHCl3); 1H NMR
C6H11NO5ꢃ0.3H2O: C, 39.47; H, 6.40; N, 7.67. Found: C, 39.25; H,
(400 MHz, CDCl3) d 1.31 (3H, dt, J = 7.3, 2.4 Hz), 1.47 (9H, d,
J = 15.0), 3.42 (1H, dt, J = 21.3, 3.9 Hz), 3.52 (1H, d, J = 3.9 Hz),
3.62 (1H, dd, J = 15.5, 10.1 Hz), 3.78 (1H, dd, J = 16.9, 15.0,
3.9 Hz), 4.18–4.32 (3H, m), 4.72 (0.5H, d, J = 5.8 Hz), 4.92 (0.5H, d,
J = 5.8 Hz), 4.95 (1H, dd, J = 11.1, 2.4 Hz); 13C NMR (100 MHz,
CDCl3) d 13.9, 13.9, 28.1, 28.2, 38.8, 39.4, 51.3, 52.1, 52.1, 52.7,
55.2, 55.4, 61.6, 61.6, 69.6, 69.7, 81.0, 81.1, 154.3, 155.0, 172.2,
172.4; IR (KBr): 3455.6, 2977.4, 1720.9, 1698.8, 1401.0, 1369.5,
1174.9, 1115.8 cmꢀ1; EI-MS (m/z): 287 (M+); HRMS Calcd for
6.25; N, 7.73.
5.21. (2S,3S,4S,5S)-1-tert-Butyl 2-ethyl 3,4,5-trihydroxypiper-
idine-1,2-dicarboxylate (L-24)
By the same procedure as for the synthesis of
(214 mg, 0.70 mmol) was obtained as a sole product from
L
-16 and
L
-17,
L
-24
-5
L
(202 mg, 0.75 mmol) after silica gel column chromatography (n-
hexane/ethyl acetate = 1:4). Mp 58–60 °C; ½a D26
ꢂ
ꢀ16.2° (c 0.90,
C
C
13H21NO6: 287.1369. Found: 287.1357; Anal. Calcd for
13H21NO6: C, 54.35; H, 7.37; N, 4.88. Found: C, 54.69; H, 7.49;
N, 4.71.
CHCl3); 1H NMR (400 MHz, CD3OD) d 1.26 (3H, t, J = 7.2 Hz), 1.45
(9H, s), 3.38–3.51 (1H, m), 3.82 (1H, d, J = 9.2 Hz), 3.91–4.04 (3H,
m), 4.16 (2H, q, J = 7.2 Hz), 4.88 (1H, d, J = 17.4 Hz); 13C NMR
(100 MHz, CD3OD) d 14.6, 28.5, 47.0, 47.8, 59.3, 60.5, 61.9, 67.6,
68.9, 69.2, 71.6, 82.0, 156.9, 157.5, 172.2; IR (KBr): 3424.0,
1732.6, 1704.6, 1394.6, 1368.2, 1254.9, 1169.7, 1086.6 cmꢀ1; EI-
MS (m/z): 305 (M+); HRMS Calcd for C13H23NO7: 305.1475. Found:
287.1461.
5.24. (2S,3S,4S,5R)-3,4,5-Trihydroxypipecolic acid(1-deoxy-5-aza-
-idopyranuronic acid,
pipecolic acid(1-deoxy-5-aza-
D
L-28) and (2S,3S,4R,5S)-3,4,5-trihydroxy-
D
-galactopyranuronic acid, -29)
L
From L-26: a mixture of L-26 (144 mg, 0.50 mmol), H2O (2.0 mL)
and H2SO4 (0.20 mL) was kept at 100 °C for 12 h. The solvent was
removed under reduced pressure and the residue was purified
twice by silica gel column chromatography (MeOH/10% NH3
aq = 50:1), then ion-exchange column (DOWEX 50W X8, eluate:
5.22. (2S,3S,4S,5S)-3,4,5-Trihydroxypipecolic acid (1-deoxy-5-
aza-D-gulopyranuronic acid,
L-25)
A mixture of
L
-24 (147 mg, 0.48 mmol) and 5 N HCl (13 mL) was
aq 0.5 N NH3) to give
28 and -29 (19 mg, 21%).
From -27: a mixture of
H2O (0.6 mL), and TFA (384 l
days. The solvent was removed under reduced pressure and the
residue was dissolved in 5 N-HCl (5 mL). The mixture was kept at
120 °C for 6 h. The solvents were removed under reduced pressure
and the residue was purified by silica gel column chromatography
L-28 (58 mg, 66%) and a 20:1 mixture ofL-
kept at 120 °C for 4 h. The solvent was removed under reduced
pressure and the residue was purified by silica gel column chroma-
L
L
L
-27 (144 mg, 0.50 mmol), THF (0.9 mL),
L, 5.0 mmol) was kept at 80 °C for 2
tography (MeOH/10% NH3 aq = 10:1) to give
L
-25 (68 mg, 80%) as a
crystal: mp 252–260 °C (decomp.); ½a D19
ꢂ
ꢀ46.4° (c 0.83, H2O); 1H
NMR (400 MHz, D2O) d 2.92 (1H, t, J = 12.1 Hz), 3.09 (1H, dd,
J = 12.1, 4.3 Hz), 3.75 (1H, d, J = 2.4 Hz), 3.91 (1H, t, J = 3.9 Hz),
4.07 (1H, ddd, J = 12.1, 4.3, 2.4 Hz), 4.22 (1H, dd, J = 4.3, 2.4 Hz);
13C NMR (100 MHz, D2O) d 42.2, 58.0, 62.8, 69.4, 69.6, 172.6
[lit.23 13C NMR (125 MHz, D2O) d 44.2, 59.9, 64.9, 71.4, 71.5,
174.8]; IR (KBr): 3258.0, 3097.3, 1644.8, 1401.2, 1096.7, 1006.2,
(MeOH/10% NH3 aq = 50:1) to give L-29 (39 mg, 44%) and a 1:3
mixture of -28 and L-29 (40 mg, 45%).
L
-28: ½a 1D8
ꢂ
L ꢀ37.6° (c 1.16, H2O); 1H NMR (400 MHz, D2O) d 3.23 (2H,
648.9 cmꢀ1
;
FAB-MS (m/z): 178 (M++1); Anal. Calcd for
m), 3.80(1H, d, J = 2.4 Hz), 3.86(1H, d, J = 2.4 Hz), 3.90(1H, t, J = 3.4 Hz),
4.14 (1H, s); 13C NMR (100 MHz, D2O) d 45.6, 58.9, 66.6, 68.2, 69.3,
172.8; IR (KBr): 3387.7, 1630.0, 1400.7, 1119.3, 1060.3, 680.8, 619.7,
473.2 cmꢀ1; FAB-MS (m/z): 178 (M++1); HRMS Calcd for C6H12NO5:
178.0715. Found: 178.0717; Anal. Calcd for C6H11NO5ꢃH2O: C, 36.92;
H, 6.71; N, 7.18. Found: C, 36.95; H, 6.62; N, 7.31.
C6H11NO5: C, 40.68; H, 6.26; N, 7.91. Found: C, 40.79; H, 6.16; N,
7.88.
5.23. (1S,4S,5S,6R)-3-tert-Butyl 4-ethy5l-hydroxy-7-oxa-3-azabi-
cyclo[4.1.0]heptane-3,4-dicarboxylate (
tert-butyl 4-ethyl 5-hydroxy-7-oxa-3-azabicyclo[4.1.0]heptane-
3,4-dicarboxylate ( -27)
L-26) and (1R,4S,5S,6S)-3-
L
-29: mp 238–247 °C (decomp.); ½a D18
ꢂ
+19.5° (c 0.75, H2O); 1H NMR
L
(400 MHz, D2O) d 2.67 (1H, t, J = 12.1 Hz), 3.31 (1H, dd, J = 12.1,
5.3 Hz), 3.52 (1H, dd, J = 9.7, 2.9 Hz), 3.66 (1H, d, J = 1.4 Hz), 3.89
(1H, ddd, J = 13.0, 9.6, 5.3 Hz), 4.29 (1H, dd, J = 2.9, 1.4 Hz); 13C NMR
(100 MHz, D2O) d 46.1, 62.8, 65.3, 69.4, 74.0, 171.9; IR (KBr): 3375.1,
3209.3, 1623.0, 1423.8, 1088.6, 688.9 cmꢀ1; FAB-MS (m/z): 178
(M++1); Anal. Calcd for C6H11NO5ꢃH2O: C, 36.92; H, 6.71; N, 7.18.
Found: C, 36.80; H, 6.67; N, 7.08.
Condition A. To a solution of L-5 (272 mg, 1.0 mmol) in CH2Cl2
(3 mL) was added a solution of mCPBA (398 mg, 1.5 mmol) in
CH2Cl2 (3 mL) at room temperature. After the mixture was stirred
at room temperature for 41 h, the reaction was quenched the by
addition of satd NaHCO3. The separated organic layer was washed
with satd NaHCO3, 10% Na2S2O3, and brine and then dried
(Na2SO4). After filtration, the filtrate was concentrated under re-
duced pressure. The residue was purified by silica gel column chro-
The
acids were synthesized from
for the -enatiomers (Schemes 13 and 14).
D
-enantiomers of 3-hydroxy- and 3,4,5-trihydroxypipecolic
D
-4 and -5 by the same methods as
D
L
matography (n-hexane/ethyl acetate = 7:3) to give more polar
(34 mg, 12%) and less polar -27 (227 mg, 79%).
Condition B. By the same procedure as for the synthesis of
-26 (581 mg, 83%) and -27 (101 mg, 14%) were obtained from
L-26
L
5.25. (2R,3R)-3-Hydroxybaikiain (D-7)
L
-21,
-5
L
L
L
½
a 2D5
ꢂ
ꢀ58.7° (c 1.0, H2O); Anal. Calcd for C6H9NO3ꢃH2O: C, 44.72;
(664 mg, 2.5 mmol) after silica gel column chromatography (n-
H, 6.88; N, 8.69. Found: C, 44.64; H, 7.06; N, 8.67.
hexane/ethyl acetate = 3:2).
L
-26: mp 82–84 °C; ½a D26
ꢂ
ꢀ74.2° (c 1.09, CHCl3); 1H NMR
5.26. (2R,3R)-1-tert-Butyl 2-ethyl 3-hydroxypiperidine-1,2-
(400 MHz, CDCl3) d 1.32 (3H, t, J = 7.2 Hz), 1.46 (9H, s), 3.23–3.37
(3H, m), 3.56 (0.5H, d, J = 8.7 Hz), 3.72 (0.5H, d, J = 8.7 Hz), 4.05
(1H, br s), 4.17–4.32 (3H, m), 4.75 (0.5H, s), 4.93 (0.5H, s); 13C
NMR (100 MHz, CDCl3) d 13.9, 27.9, 39.1, 40.3, 50.0, 50.2, 53.4,
54.0, 54.4, 55.8, 61.3, 64.5, 64.8, 81.0, 154.5, 154.8, 170.3; IR
(KBr): 3404.3, 2977.6, 1722.6, 1702.8, 1395.9, 1367.0, 1210.9,
1148.9 cmꢀ1; EI-MS (m/z): 287 (M+); HRMS Calcd for C13H21NO6:
287.1369. Found: 287.1378.
dicarboxylate (
+33.0° (c 1.15, CHCl3).
5.27. (2R,3R)-3-Hydroxypipecolic acid (
D-8)
½ ꢂ
a 2D1
D
-9)
½
a 2D8
ꢂ
ꢀ12.9° (c 1.0, H2O); Anal. Calcd for C6H11NO3ꢃ0.8H2O: C,
44.66; H, 8.00; N, 8.68. Found: C, 44.42; H, 8.03; N, 8.66.

8284
Y. Yoshimura et al. / Bioorg. Med. Chem. 16 (2008) 8273–8286
5N-HCl
120 oC, 3 h
94 %
OH
N
H
COOH
D-7
H2 / Pd-C
EtOAc
r.t., 5 h
96 %
5N-HCl
120 oC, 2h
90 %
OH
OH
N
CO2Et
N
H
COOH
Boc
-9
D
-8
D
H2SO4
H2O
VO(acac)2
100 oC, 5 h
99 %
O
(10 mol%)
TBHP
OH
CH2Cl2
r.t., 12 h
78 %
(D-22 : D-23 = 9 : 1)
N
CO2Et
Boc
OH
OH
OH
D-20
OH
HO
HO
OH
N
CO2Et
Boc
N
H
D-
COOH
N
H
COOH
D-4
D-22
23
O
CF3COCH3
Oxone®
OH
H2SO4
H2O
100 oC, h
98 %
NaHCO3
aq.Na2EDTA
CH3CN
N
CO2Et
Boc
22
23
(D- : D- = 1 : 3)
D-21
0
oC, 30 min
94 %
5N-HCl
OH
OH
120 oC, 2.5 h
quant.
HO
OH
HO
OH
N
H
COOH
N
CO2Et
K2OsO4 · 2H2O
NMO
acetone : H2O
r.t., 82 h
Boc
18
D-
D-16
33 %
+
5N-HCl
OH
OH
120 oC, 2.5 h
97 %
HO
HO
OH
OH
N
CO2Et
N
H
COOH
Boc
D-19
D-17
42 %
Scheme 13.
5.28. (2R,3S)-3-Hydroxybaikiain (
D
-11)
5.29. (2R,3S)-1-tert-Butyl 2-ethyl 3-hydroxypiperidine-1,2-
dicarboxylate ( -12)
D
½
a 2D7
ꢂ
+343.3° (c 0.74, H2O); Anal. Calcd for C6H9NO3ꢃ0.1H2O: C,
49.72; H, 6.40; N, 9.66. Found: C, 49.77; H, 6.46; N, 9.66.
½
a 2D1
+63.2° (c 0.98, CHCl3).
ꢂ

Y. Yoshimura et al. / Bioorg. Med. Chem. 16 (2008) 8273–8286
8285
5N-HCl
120 oC, 5 h
88 %
OH
N
H
COOH
11
D-
H2 / Pd-C
EtOAc
r.t., 6 h
84 %
5N-HCl
120 oC, 9.5 h
90 %
OH
CO2Et
OH
N
N
H
COOH
Boc
12
13
D-
D-
aq.H2SO4
O
100 oC, 12 h
OH
Condition A
90 %
mCPBA, CH2Cl2
r.t., 45 h, 91 %
D-28 : D-29 = 30 : 1
N
CO2Et
OH
Boc
-26 : -27 = 1 : 6
D
D
D-26
OH
HO
OH
N
CO2Et
Boc
OH
OH
HO
Condition B
-5
D
CF3COCH3
N
H
COOH
N
H
D-29
COOH
O
Oxone / NaHCO3
aq.Na2EDTA, CH3CN
OH
28
D-
0
oC, 30 min, 83 %
1) TFA,THF-H2O
80 oC, 2 days
2) 5N-HCl
N
CO2Et
-26 : -27 = 5 : 1
D
D
Boc
-27
D
120 oC, 8 h
84 %
D-28 : D-29 = 1 : 5
K2OsO4 · 2H2O
NMO
acetone : H2O = 10 : 1
r.t., 12 h
5N-HCl
OH
OH
120 oC, 7 h
89 %
OH
CO2Et
HO
HO
OH
91 %
N
N
H
COOH
Boc
-24
D
25
D-
Scheme 14.
5.30. (2R,3S)-3-Hydroxypipecolic acid (
D
-13)
5.34. (1R,4R,5S,6S)-3-tert-Butyl 4-ethyl 5-hydroxy-7-oxa-3-
azabicyclo[4.1.0]-heptane-3,4-dicarboxylate ( -20)
D
½ ꢂ
a 2D7
+51.0 ° (c 0.75, H2O); Anal. Calcd for C6H11NO3: C, 49.65; H,
7.64; N, 9.65. Found: C, 49.70; H, 7.80; N, 9.67.
½
a 2D6
+10.6° (c 1.0, CHCl3).
ꢂ
5.31. (2R,3S,4S,5S)-1-tert-Butyl 2-ethyl 3,4,5-trihydroxypiperi-
5.35. (1R,2S,3R,6S)-4-tert-Butyl 3-ethyl 2-hydroxy-7-oxa-4-
azabicyclo[4.1.0]-heptane-3,4-dicarboxylate ( -21)
dine-1,2-dicarboxylate (D-16) and (2R,3S,4R,5R)-1-tert-butyl 2-
D
ethyl 3,4,5-trihydroxypiperidine-1,2-dicarboxylate (
D-17)
½
a 2D7
ꢂ
ꢀ19.4° (c 1.17, CHCl3).
D
D
-16: ½a 2D7
ꢂ
ꢀ6.8° (c 1.42, CHCl3).
+6.0° (c 1.80), CHCl3).
-17: ½a 2D7
ꢂ
5.36. (2R,3S,4R,5S)-3,4,5-Trihydroxypipecolic acid(1-deoxy-5-aza-
-altropyran-uronic acid, -22) and (2R,3S,4S,5R)-3,4,5-trihydroxy
L
D
5.32. (2R,3S,4S,5S)-3,4,5-Trihydroxypipecolic acid (1-deoxy-5-
aza- -allopyran-uronic acid, -18)
pipecolic acid(1-deoxy-5-aza- -glucopyranuronic acid, -23)
L
D
L
D
D
-22: ½a 2D6
+11.0° (c 0.90, H2O); Anal. Calcd for C6H11NO5ꢃ0.5H2O:
ꢂ
½
a 2D7
ꢂ
+29.4° (c 1.1, H2O); Anal. Calcd for C6H11NO5ꢃ0.2H2O: C,
C, 38.71; H, 6.50; N, 7.52. Found: C, 39.01; H, 6.70; N, 7.22.
39.87; H, 6.36; N, 7.75. Found: C, 40.00; H, 6.20; N, 7.68.
D
-23: ½a 2D6
ꢀ18.1° (c 0.78, H2O); Anal. Calcd for C6H11NO5
ꢂ
ꢃ1.5H2O: C, 35.29; H, 6.91; N, 6.86. Found: C, 35.25; H, 6.79; N, 6.95.
5.33. (2R,3S,4R,5R)-3,4,5-Trihydroxypipecolic acid (1-deoxy-5-
aza-L-mannopyran-uronic acid, D-19)
5.37. (2R,3R,4R,5R)-1-tert-Butyl 2-ethyl 3,4,5-
trihydroxypiperidine-1,2-dicarboxylate (
D-24)
½
a 2D6
ꢂ
ꢀ30.0° (c 1.1, H2O); Anal. Calcd for C6H11NO5ꢃ0.2H2O: C,
39.87; H, 6.36; N, 7.75. Found: C, 39.69; H, 6.46; N, 7.62.
½ ꢂ
a 2D6
+16.8° (c 1.06, CHCl3).

8286
Y. Yoshimura et al. / Bioorg. Med. Chem. 16 (2008) 8273–8286
2. (a) Kato, A.; Kato, N.; Kano, E.; Adachi, I.; Ikeda, K.; Yu, L.; Okamoto, T.; Banba,
Y.; Ouchi, H.; Takahata, H.; Asano, N. J. Med. Chem. 2005, 48, 2036; (b) Ruiz, M.;
Ruanova, T. M.; Blanco, O.; Nún¨ez, F.; Pato, C.; Ojea, V. J. Org. Chem. 2008, 73,
2240.
3. Mori, K.; Kondo, T.; Kamiyama, Y.; Kano, Y.; Tominaga, K. Cancer Chemother.
Pharmacol. 2003, 51, 403.
4. (a) Bello, I. C.; Dorling, P.; Fellows, L.; Winchester, B. FEBS Lett. 1984, 176, 61; (b)
Tong, M. K.; Blumenthal, E. M.; Ganem, B. Tetrahedron Lett. 1990, 31, 1683; (c)
Tsuruoka, T.; Fukuyasu, H.; Ishii, M.; Usui, T.; Shibahara, S.; Inouye, S. J. Antibiot.
1996, 49, 155.
5. (a) Sugisaki, C. H.; Caroll, P. J.; Correia, C. R. D. Tetrahedron Lett. 1998, 39, 3413;
(b) Hanessian, S.; McNaughton-Smith, G.; Lombart, H.-G.; Lubell, W. D.
Tetrahedron 1997, 53, 12789.
5.38. (2R,3R,4R,5R)-3,4,5-Trihydroxypipecolic acid (1-deoxy-5-
aza- -gulopyranuronic acid, -25)
L
D
½
a 1D9
ꢂ
+46.4° (c 1.23, H2O) [lit.16
½
a 2D5
ꢂ
ꢀ5.9° 2.2 (c 1.25, variable,
H2O)]; Anal. Calcd for C6H11NO5: C, 40.68; H, 6.26; N, 7.91. Found:
C, 40.70; H, 6.18; N, 7.92.
5.39. (1R,4R,5R,6S)-3-tert-butyl 4-ethy5l-hydroxy-7-oxa-3-azabi-
cyclo[4.1.0]-heptane-3,4-dicarboxylate (
3-tert-butyl 4-ethyl 5-hydroxy-7-oxa-3-azabicyclo[4.1.0]heptane-
3,4-dicarboxylate ( -27)
D-26) and (1S,4R,5R,6R)-
6. Kusano, G.; Ogawa, H.; Takahashi, A.; Nozoe, S.; Yokoyama, K. Chem. Pharm.
Bull. 1987, 35, 3482.
D
7. Synthesis of 3-hydroxypipecoli acid was reported: (a) Kumar, P.; Bodas, M. S. J.
Org. Chem. 2005, 70, 360; (b) Scott, J. D.; Williams, R. M. Tetrahedron Lett. 2000,
41, 8413; (c) Liang, N.; Datta, A. J. Org. Chem. 2005, 70, 10182.
8. A part of this work has appeared: Ohara, C.; Takahashi, R.; Miyagawa, T.;
Yoshimura, Y.; Kato, A.; Adachi, I.; Takahata, H. Bioorg. Med. Chem. Lett. 2008,
18, 1810.
D
D
-26. ½a 2D5
ꢂ
+73.4° (c 1.23, CHCl3).
+50.7° (c 0.90, CHCl3).
-27. ½a 2D0
ꢂ
5.40. (2R,3R,4R,5S)-3,4,5-Trihydroxypipecolic acid(1-deoxy-5-
aza- -idopyranuronic acid, -28) and (2R,3R,4S,5R)-3,4,5-trihydr-
oxypipecolic acid(1-deoxy-5-aza- -galactopyranuronic acid, -29)
9. Simone, S.; Gunter, H. Eur. J. Org. Chem. 1999, 2515.
D
D
10. Compound
4
was converted to
a
known 2,3-trans-2-hydroxy-3-
L
D
hydroxymethyl-piperidine in three steps: (1) H2, Pd–C, EtOAc; 99%. (2)
LiAlH4, THF; 79%. (3) 5 N HCl, MeOH; 88%.
D
D
-28: ½a 1D8
ꢂ
+35.7° (c 0.87, H2O) [lit.28
ꢀ20.0° (c 0.77, H2O); Anal. Calcd for C6H11NO5ꢃH2O:
½
a 2D5
+39.7° (c 1.1, H2O)]
ꢂ
11. The ee of
Ind., Ltd, Japan after deacetylation followed by replacing a Boc to a tosyl group.
Similarly, the ee of -6 was determined by HPLC after replacing a Boc to a tosyl
group (see Section 5).
12. The ees of -5 and
described in Ref. 11.
13. Both -16 and -17 were converted to 1-deoxymammonojirimycin and 1-
D-6 was determined by HPLC using CHIRALPAK IA, Daicel Chemical
-29: ½a 1D9
ꢂ
L
C, 36.92; H, 6.71; N, 7.18. Found: C, 37.09; H, 6.65; N, 7.16.
L
D-4 (D-10) were determined by the same method as
5.41. Evaluation of inhibitory activities against glycosidases
L
L
deoxyallonojirimycin in two steps (1) LiAlH4, THF; (2) 5 N HCl, MeOH and
compared their instrumental data with those reported (see Ref. 2).
14. (a) Park, K. H.; Yoon, Y. J.; Lee, S. G. J. Chem. Soc., Perkin Trans. 1 1994, 2621; (b)
Fleet, G. W. J.; Ramsden, N. G.; Witty, D. R. Tetrahedron 1989, 45, 327; (c) Fleet,
G. W. J.; Fellows, L. E.; Smith, P. W. Tetrahedron 1987, 43, 979.
15. (a) Tsimilaza, A.; Tite, T.; Boutefnouchet, S.; Lallemand, M.; Tillequin, F.;
Husson, H. Tetrahedron Asymmetry 2007, 18, 1585; (b) Bashyal, B. P.; Choe, H.-
F.; Fleet, G. W. J. Tetrahedron Lett. 1986, 27, 3205; (c) Ref. 14c.
16. Park, K. H. Bull. Korean Chem. Soc. 1995, 16, 985.
17. (a) Hoveyda, A. H.; Evans, D. A.; Fu, G. C. Chem. Rev. 1993, 93, 1307; (b)
Chamberlain, P.; Roberts, M. L.; Whitham, G. H. J. Chem. Soc. B 1970, 1374.
18. (a) Torres, C. R.; Hart, G. W. J. Biol. Chem. 1984, 259, 3308; (b) Wells, L.;
Vosseller, K.; Hart, G. W. Science 2001, 291, 2376; (c) Hanover, J. A. FASEB J.
2001, 15, 1865.
The enzymes b-glucuronidase (from bovine liver, assayed at pH
5.0; from E. coli, assayed at pH 6.8), b-N-acetylglucosaminidase
(from bovine kidney, pH 4.5; from human placenta, assayed at pH
4.5),
a-N-acetylgalactosaminidase (from chicken liver, pH 4.0), p-
nitrophenyl glycosides were purchased from Sigma Chemical Co.
Glycosidase activities were determined using an appropriate p-
nitrophenyl glycoside as substrate at optimum pH of each enzyme.
The reaction mixture (1 mL) contained 2 mM of the substrate and
the appropriate amount of enzyme. The reaction was stopped by
adding 2 mL of 400 mM Na2CO3. The released p-nitrophenol was
measured spectrometrically at 400 nm. Enzyme inhibition modes
and Ki values were determined from Lineweaver–Burk plots.
19. Goldberg, H. J.; Whiteside, C. I.; Hart, G. W.; Fantus, I. G. Endocrinology 2006,
147, 222.
20. Hart, G. W. Annu. Rev. Biochem. 1997, 66, 315.
21. Vosseller, K.; Wells, L.; Lane, M. D.; Hart, G. W. Proc. Natl. Acad. Sci. U.S.A. 2002,
99, 5313.
22. Mar, B. L.; Vocadlo, D. J.; Knapp, S.; Triggs-Raine, B. L.; Withers, S. G.; James, M.
N. G. J. Biol. Chem. 2001, 276, 10330.
23. Gao, Y.; Wells, L.; Comer, F. I.; Parker, G. J.; Hart, G. W. J. Biol. Chem. 2001, 276,
9838.
24. (a) Knapp, S.; Abdo, M.; Ajayi, K.; Huhn, R. A.; Emge, T. J.; Kim, E. J.; Hanover, J.
A. Org. Lett. 2006, 9, 2321; (b) Whitworth, G. E.; Macauley, M. S.; Stubbs, K. A.;
Dennis, R. J.; Taylor, E. J.; Davies, G. J.; Greig, I. R.; Vocadlo, D. J. J. Am. Chem. Soc.
2006, 129, 635; (c) Shanmugasundaram, B.; Debowski, A. W.; Dennis, A. W.;
Dennis, R. J.; Davis, G. J.; Vocadlo, D. J.; Vasella, A. Chem. Commun. 2006, 4372.
25. Gandia, D.; Abigerges, D.; Armand, J.-P.; Chabot, G.; Da Costa, L.; De Forni, M.;
Herait, P. J. Clin. Oncol. 1993, 11, 196.
26. Kaneda, N.; Nagata, H.; Furuta, T.; Yokokura, T. Cancer Res. 1990, 50, 1715.
27. Horikawa, M.; Busch-Petersen, J.; Corey, E. J. Tetrahedron Lett. 1999, 40, 3843.
28. (a) Lee, B. W.; Jeong, I.-Y.; Yang, M. K.; Choi, S. K.; Park, K. H. Synthesis 2000,
1305; (b) Bashyal, B. P.; Chow, H.-F.; Fleet, G. W. J. Tetrahedron Lett. 1986, 27,
3205.
Acknowledgments
This work was supported, in part, by a Grant-in-Aid for Scien-
tific Research (No. 18590012) (H.T.) and by the High Technology
Research Program from the Ministry of Education, Culture, Sports,
Science and Technology of Japan.
References and notes
1. (a)Iminosugars as Glycosidase Inhibitors: Nojirimycin and Beyond; Stutz, A. E., Ed.;
Wiley-VCH: Weinheim, 1999; (b) Asano, N.; Nash, R. J.; Molyneux, R. J.; Fleet, G.
W. J. Tetrahedron Asymmetry 2000, 11, 1645; (c) Watson, A. A.; Fleet, G. W. J.;
Asano, N.; Molyneux, R. J.; Nash, R. J. Phytochemistry 2001, 56, 265; (d) Martin,
O. R., Compain, P., Eds. Curr. Top. Med. Chem. 2003, 3.; (e) Asano, N. Glycobiology
2003, 13, 93R.