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R. Watanabe et al. / Tetrahedron Letters 43 (2002) 6501–6504
The molecular formula of 1 was determined to be
C15H26N2O6 by ESI MS (m/z 331.1870, calcd for
C15H27N2O6 [M+H]+ 331.1869). The NMR data for 1
1
are summarized in Table 1. The H NMR, 13C NMR,
and HMQC spectra of 1 showed the presence of one
methyl carbon (lc 12.8), eight methylene carbons, two
methine carbons (lc 54.0, 54.5) connected to hetero
atoms, and four carbonyl carbons (lc 173.3, 174.1,
176.9, 213.7). A detailed analysis of the COSY spec-
trum of 1 allowed three partial structures, C-2 to C-5,
C-7 to C-10, and C-2% to C-4% (Fig. 2). The remaining
connectivities of 1 were clarified by the HMBC correla-
tions H-2/C-1 and C-5%, H-5/C-6, H-7/C-6, H-2%/C-1%,
and H-4%/C-5%. Thus, the gross structure of N-g-glu-
tamyl boletine was determined to be a dipeptide con-
sisting of glutamic acid and a new amino acid, which
was named boletine, as shown in 1.
Scheme 1. Hydrolysis of N-g-glutamyl boletine (1).
the carbon adjacent to the carboxyl group in 4 could
not be determined.
To confirm the absolute stereochemistry of N-g-glu-
tamyl boletine (1) and to obtain a greater supply for
further biological studies, both diastereomers, (2S, 2%S)-
and (2R, 2%S)-dipeptides 11 and 12, were synthesized
(Scheme 2). The synthesis of 1 started from commer-
The absolute stereostructure of 1 was elucidated as
follows. Acidic hydrolysis of 1 followed by separation
using reversed-phase HPLC gave glutamic acid (3) and
cyclic amino acid 4 (Scheme 1). The absolute configura-
tion of the glutamic acid (3) was determined to be L by
chiral HPLC analysis.3 The gross structure of cyclic
amino acid 4 was determined by analysis of COSY,
HMQC and HMBC spectra, but the stereochemistry of
cially available Boc- -Asp-OBn (5). Compound 5 was
L
converted into a mixed anhydride and then subjected to
reduction using sodium borohydride to give alcohol 6
in 69% yield.4 Dess–Martin oxidation of alcohol 6 gave
aldehyde 7 in 79% yield. The Horner–Wadsworth–
Emmons reaction of aldehyde 7 with b-ketophospho-
nate gave a,b-unsaturated ketone 8 in 77% yield.5,6 The
geometry of the olefin in 8 was confirmed to be E based
on the magnitude of the coupling constant between
vinyl protons (16.0 Hz). Acidic deprotection of the Boc
Table 1. NMR data for N-g-glutamyl boletine (1) in
CD3OD
group in 8 followed by condensation with Z-L-Glu-OBn
13Ca
1Hb
HMBCc
(9) gave dipeptide 10 in 53% yield. Finally, hydrogena-
tion of 10 using Pd(OH)2 led to deprotection of the Z
and Bn groups and reduction of the olefin to give (2S,
2%S)-dipeptide 11 in quantitative yield. Similarly, (2R,
Atom
1
2
3
4
5
6
7
8
176.9 s
54.0 d
31.4 t
20.3 t
41.0 t
213.7 s
41.7 t
26.8 t
22.4 t
12.8 q
173.3 s
54.5 d
27.0 t
H-2, 3
H-3, 4
H-2, 5
H-2, 3, 5
H-3
H-5, 7, 8
H-8, 9
H-7, 9, 10
H-7, 8, 10
H-8, 9
H-2%, 3%
H-3%, 4%
H-2%, 4%
4.23 m 1H
1.62 m 2H
1.84 m 2H
2.51 m 2H
2%S)-dipeptide 12 was synthesized from Boc-D-Asp-OBn
(ent-5) in 27% yield in six steps. Synthetic (2S, 2%S)-
dipeptide 11 was identical to natural 1 with respect to
TLC analysis and spectroscopic data (1H and 13C
NMR, and optical rotation).7 Thus, the absolute
stereostructure in N-g-glutamyl boletine (1) was shown
to be 2S and 2%S. Synthetic N-g-glutamyl boletine (11)
also exhibited moderate antibacterial activity against R.
salexigens (9 mm, 1 mg/disk) as with natural products.
It is interesting to note that 2R-isomer 12 did not show
any antibacterial activity.
2.46 t (6.7) 2H
1.51 tt (6.7, 6.9) 2H
1.31 tq (6.9, 6.9) 2H
0.92 t (6.9) 3H
9
10
1%
2%
3%a
3%b
4%
5%
3.64 m 1H
2.10 m 1H
2.23 m 1H
2.52 m 2H
32.5 t
174.1 s
H-2%, 3%
H-2, 3%, 4%
2-Butyl-1-azacyclohexene iminium salt (2) showed an
ion peak at m/z 140 [M+] in its FAB MS spectrum. The
a Recorded at 200 MHz. Multiplicity was based on the HMQC
spectrum.
1
NMR data for 2 are summarized in Table 2. The H
NMR, 13C NMR, and HMQC spectra of 2 showed the
presence of one methyl carbon (lc 12.6), seven methyl-
ene carbons, and one carbonyl carbon (lc 192.3). Analy-
sis of the COSY, HMQC, HMBC spectra of 2 allowed
us to construct its entire framework (Fig. 3). Treatment
b Recorded at 800 MHz. Coupling constants (Hz) are in parentheses.
c Based on the correlation from each carbon atom.
1
of 2 with triethylamine gave an imine, the H NMR
data of which coincided with those of synthetic 2-butyl-
1-azacyclohexene previously reported.8 Thus, the struc-
ture was confirmed to be as shown in Fig. 3.
The biosynthesis of piperidine alkaloids has been well
studied. Experiments using labeled precursors have
established that a large group of piperidine alkaloids
can be derived from lysine or acetic acid.9 For example,
Figure 2. Gross structure of N-g-glutamyl boletine (1).