Tetrahedron Letters 53 (2012) 1227–1230
Contents lists available at SciVerse ScienceDirect
Tetrahedron Letters
journal homepage: www.elsevier.com/locate/tetlet
Leucomidines A–C, novel alkaloids from Leuconotis griffithii
Midori Motegi a, Alfarius Eko Nugroho a, Yusuke Hirasawa a, Takashi Arai a, A. Hamid A. Hadi b,
Hiroshi Morita a,⇑
a
b
Faculty of Pharmaceutical Sciences, Hoshi University, Ebara 2-4-41, Shinagawa-ku, Tokyo 142-8501, Japan
Department of Chemistry, Faculty of Science, University of Malaya, Kuala Lumpur 50603, Malaysia
a r t i c l e
i n f o
Article history:
Received 24 November 2011
Revised 20 December 2011
Accepted 26 December 2011
Available online 2 January 2012
a b s t r a c t
Two novel indole alkaloids and new quinazoline alkaloids, namely leucomidines A–C (1–3), were isolated
from the barks of Leuconotis griffithii. The structures including the absolute configuration were elucidated
on the basis of spectroscopic data, chemical means, and CD calculations. The quinazoline alkaloid, 3, is
proposed to be derived from the same biogenetic precursor as the indole alkaloids 1 and 2.
Ó 2012 Elsevier Ltd. All rights reserved.
Keywords:
Indole alkaloid
Quinazoline alkaloid
Leuconotis griffithii
Apocynaceae
Leuconotis griffithii (Retz.) Gardner ex Thwaites is a member of
the Apocynaceae family in Malaysia and Indonesia.1 The species
of Leuconotis have been known to produce monoterpene indole
alkaloids,2,3 whose skeletons are similar to those found in Alstonia and Kopsia species.4 The species of Alstonia, Kopsia, Hunteria,
Tabernaemontana, and Leuconotis have been known to produce
various alkaloids depending on the area where the plants were
distributed.2–7 Recently, we isolated a new bisindole alkaloid, bisleuconothine A consisting of an eburnane–aspidosperma-type
skeleton from the barks of L. griffithii.5 In our continuing search
for structurally and biogenetically interesting alkaloids from tropical plants, we isolated three novel alkaloids, leucomidines A–C
( 1–3), together with leuconolam4 and O-methylleuconolam.4 In
this Letter, the isolation and structure elucidation of 1–3 are
described.
Leucomidine A (1),8,9 yellowish amorphous solid, ½a24
+69
D
(c 0.3, MeOH), showed molecular formula, C19H20N2O3, which
was determined by HRESIMS [m/z 347.1380 (M+Na)+, D
+0.1 mmu]. IR absorption bands were characteristic of amino or
hydroxyl (3390 cm1) group and two carbonyl (1800 and
1730 cm1) groups. 1H and 13C NMR data (Table 1) suggested the
presence of six sp3 methylenes, a methyl, two sp3 quaternary
carbons, four sp2 methines, and six sp2 quaternary carbons.
⇑ Corresponding author. Tel./fax: +81 354985778.
E-mail address: moritah@hoshi.ac.jp (H. Morita).
0040-4039/$ - see front matter Ó 2012 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetlet.2011.12.116
The gross structure of 1 was deduced from extensive analyses of
the two-dimensional NMR data, including the 1H–1H COSY, HSQC,
and HMBC spectra in CD3OD (Fig. 1). The 1H–1H COSY and HSQC
spectra revealed connectivity of four partial structures a (C-9–C12), b (C-3, C-14–C-15), c (C-16–C-17), and d (C-18–C-19), as
1228
M. Motegi et al. / Tetrahedron Letters 53 (2012) 1227–1230
Table 1
1
H (600 MHz) and
13
C (150 MHz) NMR data of leucomidines A–C (1–3)
1a
2b
dH (J, Hz)
2
3a
3b
5
6
7
8
9
10
11
12
13
14a
14b
15a
15b
16a
16b
17a
17b
18
19a
19b
20
21
OMe
a
b
dC
2.73 (1H, ddd, 13.1, 13.0, 4.2)
4.25 (1H, dd, 13.1, 4.9)
6.98
6.98
7.11
7.35
(1H,
(1H,
(1H,
(1H,
br s)
d, 6.1)
ddd, 8.1, 6.1, 2.1)
d, 8.1)
1.66
1.53
1.74
1.73
2.91
(1H,
(1H,
(1H,
(1H,
(2H,
m)
br dd, 13.1, 3.1)
dd, 9.1, 3.1)
br s)
m)
2.05
2.02
0.92
1.46
1.32
(1H,
(1H,
(3H,
(1H,
(1H,
dt, 7.6, 3.2)
ddd, 18.4, 13.9 10.1)
t, 7.6)
dq, 14.9, 7.6)
dq, 14.9, 7.6)
141.8
40.7
154.1
161.1
103.6
125.4
117.6
121.5
123.0
112.7
138.3
20.7
26.6
19.9
29.5
7.6
23.1
42.7
96.1
3b
dH (J, Hz)
dC
2.98 (1H, ddd, 12.1, 7.9, 3.8)
4.30 (1H, br d, 12.1)
7.68
7.16
7.27
7.49
(1H,
(1H,
(1H,
(1H,
d, 8.2)
br t, 8.2)
br t, 8.2)
d, 8.2)
1.60
1.69
1.59
1.76
2.60
2.74
2.18
(1H,
(1H,
(1H,
(1H,
(1H,
(1H,
(2H,
m)
m)
m)
br d, 11.7)
m)
m)
t, 8.6)
0.58 (3H, t, 7.6)
0.67 (1H, dq, 14.5, 7.6)
1.12 (1H, dq, 14.5, 7.6)
4.36 (1H, s)
3.78 (3H, s)
175.3
40.1
162.8
127.6
135.2
128.2
121.4
122.5
124.8
114.1
142.5
21.2
30.1
29.2
32.7
7.6
23.7
40.2
63.8
52.4
dH (J, Hz)
dC
175.4
49.0
4.01 (1H, m)
4.10 (1H, m)
8.17
7.47
7.78
7.64
(1H,
(1H,
(1H,
(1H,
dd, 8.2, 1.0)
ddd, 8.2, 6.9, 1.3)
ddd, 8.3, 6.9, 1.0)
br d 8.3)
2.05 (2H, m)
1.85
1.99
2.30
2.54
2.12
2.27
0.88
1.76
2.11
(1H,
(1H,
(1H,
(1H,
(1H,
(1H,
(3H,
(1H,
(1H,
ddd, 12.4, 6.9,
12.4, 8.3, 4.1)
ddd, 15.8, 8.9,
ddd, 15.8, 8.9,
ddd, 14.8, 8.9,
ddd, 14.8, 8.9,
t, 7.6)
dq, 14.8, 7.6)
dq, 14.8, 7.6)
4.5)
29.3
6.1)
6.1)
6.1)
6.1)
29.1
3.41 (3H, s)
In CD3OD.
In CD3OD/CDCl3.
Figure 1. Selected 2D NMR correlations for leucomidine A (1).
Figure 3. Selected 2D NMR correlations for leucomidine B (2).
Scheme 1. Reduction of leucomidine A (1) by LiAlH4.
Figure 2. Selected NOESY correlations for leucomidine A (1, 20R⁄, 21S⁄).
shown in Figure 1. The presence of an indole ring (C-2, C-7–C-13,
and N-1) and the connectivity of partial structure c and the indole
ring were revealed by the HMBC correlations of H-9 to C-7 and
164.0
120.5
126.8
127.5
135.0
127.7
148.3
19.7
Figure 4. Selected NOESY correlations for leucomidine B (2).
35.8
8.4
32.6
44.6
160.0
51.4
M. Motegi et al. / Tetrahedron Letters 53 (2012) 1227–1230
Figure 5. Selected 2D NMR correlations for leucomidine C (3).
Figure 6. Calculated and experimental CD spectra of 1 (left scale) and 2 (right
scale).
C-13, H-10 and H-12 to C-8, H2-16 to C-7, and H2-17 to C-2. HMBC
correlations of H3-18 to C-20, H2-19 to C-15, C-17, and C-21 established the connections among C-15, C-17, C-19, and C-21 through
C-20. The HMBC cross-peaks of H2-3 to C-5 and C-21 and the
chemical shift of C-3 (dC 40.7), C-5 (dC 154.1) and C-21 (dC 96.1)
suggested the connection among C-3, C-5 and C-21 through a
nitrogen atom. Based on the chemical shift of C-21 and the
1229
remaining carbonyl carbon (dC 161.1), the structure of 1 was assumed to be as shown in Figure 1.
To clarify the structure, 1 was reduced by LiAlH4 (Scheme 1) to
obtain 4 and 5 whose structures further support the proposed
structure of 1 as shown in Figure 1.10
The lack of proton on C-5 and C-6 complicates the determination of the relative configuration of C-20 and C-21. Fortunately,
the 20R⁄,21R⁄ and the 20R⁄,21S⁄ isomers have different stable conformation of the piperidine unit (boat and chair, respectively). The
NOESY correlations and coupling constant data suggested that the
piperidine unit in 1 took a chair conformation (Fig. 2), thus the relative configuration of 1 was deduced to be 20R⁄, 21S⁄.
Leucomidine B (2),11 yellowish amorphous solid, ½a26
D 18 (c
0.3, CHCl3), showed molecular formula, C20H24N2O3, which was
determined by HRESIMS [m/z 341.1840 (M+H)+, D 2.0 mmu]. IR
absorption bands were characteristic of amino or hydroxyl
(3400 cm1) and two carbonyl (1740 and 1670 cm1) groups. 1H
and 13C NMR data (Table 1) suggested the presence of a sp3
methine, six sp3 methylenes, two methyls, a sp3 quaternary carbon,
four sp2 methines, and six sp2 quaternary carbons.
The planar structure of 2 was deduced from extensive analyses
of the two-dimensional NMR data (1H–1H COSY, HSQC, and HMBC
spectra, Fig. 3). The 1H–1H COSY and HSQC spectra revealed four
partial structures a (C-9–C-12), b (C-3, C-14–C-15), c (C-18–C19), and d (C-16–C-17), as shown in Figure 3. HMBC correlations
of H3-18 to C-20, H2-19 to C-17 and C-21, and H2-17 to C-15 suggested the connection of C-15, C-17, C-19, and C-21 through C-20.
The presence of a piperidine ring system was deduced from the
chemical shift of C-3 (dC 40.1) and C-21 (dC 63.8), and from the
HMBC correlations of H-21 to C-3. HMBC correlations of H2-17
and H3-OMe to C-2 revealed the connection of a methoxy carbonyl
group at C-16. Finally, the HMBC cross-peaks of H-21 to C-5, C-6,
and C-7 were used to deduce the presence of a pyrolone ring system bridging the indole and the piperidine units.
The relative configuration of 2 was determined by NOESY spectral data (Fig. 4). The b-orientation of H-21 and C-17 was suggested
by NOESY correlations of H2-17/H-21.
Leucomidine C (3),12 yellowish amorphous solid, ½a26
D +32 (c 0.3,
CHCl3), showed molecular formula, C18H22N2O3, which was
Figure 7. Plausible biogenetic pathway of 1–3.
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M. Motegi et al. / Tetrahedron Letters 53 (2012) 1227–1230
determined by HRESIMS [m/z 315.1685 (M+H)+, D 1.8 mmu]. IR
absorption bands suggested the presence of two carbonyl (1740
and 1680 cm1) groups. 1H and 13C NMR data (Table 1) of 3 showed
characteristic signals13 of a 2,3-disubstituted 4(3H)-quinazolinones ring [dC 164.0 (C-7), 120.5 (C-8), 126.8 (C-9), 127.5 (C-10),
135.0 (C-11), 127.7 (C-12), 148.3 (C-13) and 160.0 (C-21); dH 8.17
(H-9), 7.47 (H-10), 7.78 (H-11) and 7.64 (H-12)]. In addition to
the quinazolinone ring, the 1H and 13C NMR data suggested the
presence of one sp3 quaternary carbon, six sp3 methylenes, two
methyls, and one sp2 quaternary carbon.
Analyses of the 1H–1H COSY and HSQC spectra (Fig. 5) revealed
the presence of partial structure b (C-3, C-14–C-15), c (C-18–C-19),
and d (C-16–C-17), in addition to partial structure a (C-9–C-12)
which is part of the quinazoline ring. HMBC correlations of H3-18
to C-20, H2-19 to C-15, C-17, and C-21 suggested the connection
of C-15, C-17, C-19, and C-21 through C-20 while the HMBC correlation of H2-3 to C-21 and the chemical shift of C-3 indicated that
C-3 is connected to N-4. Finally, the presence of a methoxy carbonyl group at C-16 was deduced from the HMBC correlation of
H2-17 and H3-OMe to C-2. Thus, the structure of 3 was deduced
to be as shown in Fig. 5.
The absolute configuration of 1 and 2 were determined by comparing the calculated14 and the experimental CD spectra. As can be
seen in Fig. 6, the calculated CD spectra of the 20R, 21S isomer of
both compounds are similar to the experimental one. Thus, the
absolute configurations of 1 and 2 were assigned as 20R, 21S.
Biogenetically,1–3 are assumed to be derived from leuconolam4
(Fig. 7). Oxidation of C-6 and C-7 of leuconolam may give precursor
A. 1 might be produced by ring closures of A, whereas hydrolysis of
A followed by ring closures and reduction of OH on C-21 to H
might give 2. Further oxidation of A followed by hydrolysis will
give B, and 3 may be obtained from intermediate C after several
steps. Considering the biogenetic route, though the absolute configuration of 3 could not be assigned by using CD calculation, the
absolute configuration at C-20 of 3 is proposed to be R.
Leucomidines A–C (1–3) were tested for cytotoxic activity
against HL-60 cell line.15 All three compounds were found to be
inactive (IC50 >100 lM).
Acknowledgments
This work was partly supported by a Grant-in-Aid for Scientific
Research from the Ministry of Education, Culture, Sports, Science,
and Technology of Japan, a grant from the Open Research Center
Project and Takeda Science Foundation, and also a grant from
HIR UM-MOHE (F000009-21001).
References and notes
1. Whitmore, T. C. In Tree Flora Malaysia; Whitmore, T. C., Ed.; Academic Press:
London, 1972; Vol. 2, pp 3–24.
2. Gan, C. Y.; Robinson, W. T.; Etoh, T.; Hayashi, M.; Komiyama, K.; Kam, T. S. Org.
Lett. 2009, 11, 3962–3965.
3. Gan, C. Y.; Kam, T. S. Tetrahedron Lett. 2009, 50, 1059–1061.
4. Goh, S. H.; Ali, A. R. M.; Wong, W. H. Tetrahedron 1989, 45, 7899–7920.
5. Hirasawa, Y.; Shoji, T.; Arai, T.; Nugroho, A. E.; Deguchi, J.; Hosoya, T.;
Uchiyama, N.; Goda, Y.; Awang, K.; Hadi, A. H. A.; Shiro, M.; Morita, H. Bioorg.
Med. Chem. Lett. 2010, 20, 2021–2024.
6. Nugroho, A. E.; Hirasawa, Y.; Kawahara, N.; Goda, Y.; Awang, K.; Hadi, A. H. A.;
Morita, H. J. Nat. Prod. 2009, 72, 1502–1506.
7. Hirasawa, Y.; Miyama, S.; Hosoya, T.; Koyama, K.; Rahman, A.; Kusumawati, I.;
Zaini, N. C.; Morita, H. Org. Lett. 2009, 11, 5718–5721.
8. The barks of L. griffithii collected at Malaysia, were extracted with MeOH, and
part (17 g) of the extract was treated with 3% tartaric acid (pH 2) and then
partitioned with EtOAc. The EtOAc fraction was subjected to a silica gel column
(hexane/EtOAc, CHCl3/MeOH) to give 27 fractions. Fraction containing 1 was
further separated using a silica gel column (hexane/EtOAc, toluene/EtOAc) to
give leucomidine A (1, 3.1 mg, 0.005%). While fraction containing 2 and 3 was
further separated using an ODS silica gel column (MeOH/H2O) and ODS HPLC
(MeOH/H2O) to give leucomidines B (2, 0.2 mg, 0.0003%) and C (3, 0.6 mg,
0.0009%).
9. Leucomidine A (1): yellowish amorphous solid; ½a24
D +69 (c 0.3, MeOH); IR
(KBr) mmax 3390, 2950, 1800, and 1730 cm1; UV (MeOH) kmax 301 (e 2200),
250 (8100), and 216 (34100) nm; CD (MeOH) kmax 287 (De +0.64), 277 (0), 227
(+15.94), 218 (0), and 209 (8.97) nm; 1H and 13C NMR (Table 1); ESIMS (pos.)
m/z 347 (M+Na)+; HRESITOFMS m/z 347.1380 (M+Na)+, calcd for C19H20N2O3Na
347.1379.
10. To a solution of 1 (2.0 mg) in THF (1 ml) was added LiAlH4 (5 mg). The mixture
was allowed to stand at 80 °C for 3 h. After cooling, the mixture was
partitioned between H2O and CHCl3. CHCl3 soluble materials were applied to
a silica gel column (CHCl3/MeOH, 1:0?0:1) to give compounds 4 (0.7 mg) and
5 (0.6 mg). 4, 1H NMR (CD3OD, 600 MHz) 7.51 (d, 7.6; H-9), 7.29 (d, 7.6; H-12),
7.06 (t, 7.6; H-11), 7.04 (t, 7.6; H-10), 3.75 (br. s; H-21), 3.63 (m; H2-6), 3.47
(m; H-3a), 3.34 (m; H2-5), 2.93 (dd, 17.2, 7.2; H-16a), 2.79 (ddd, 17.2, 11.0, 7.2;
H-16b), 2.68 (m; H-3b), 2.64 (m; H-17a), 1.96 (m; H-14a), 1.80 (br. d, 15.2; H15a), 1.77 (m; H-14b), 1.63 (br. t, 12.7; H-15b), 1.55 (dd, 13.0, 7.2; H-17b), 1.23
(dq, 14.1, 7.6; H-19a), 1.08 (dq, 14.1, 7.6; H-19b), and 0.80 (t, 7.6; H3-18); 13C
NMR (CD3OD, 150 MHz) 138.4 (C-2), 138.3 (C-13), 130.3 (C-8), 122.1 (C-11),
120.6 (C-10), 118.6 (C-9), 111.9 (C-12), 103.6 (C-7), 66.3 (C-21), 58.7 (C-6), 57.8
(C-5), 53.8 (C-3), 38.1 (C-20), 34.6 (C-15), 30.7 (C-19), 24.9 (C-17), 21.3 (C-14),
20.8 (C-16), and 7.9 (C-18); ESIMS (pos.) m/z 299 (M+H)+. 5, 1H NMR (CD3OD,
600 MHz) 7.99 (d, 7.2; H-9), 7.41 (d, 7.2; H-12), 7.24 (td, 7.2, 1.4; H-11), 7.21
(td, 7.2, 1.4; H-10), 3.85 (dd, 15.8, 6.2; H-3a), 3.69 (ddd, 15.8, 10.7, 6.2; H-3b),
3.09 (ddd, 17.9, 12.7, 5.5; H-16a), 2.96 (ddd, 17.9, 5.2, 1.4; H-16b), 2.20 (ddd,
12.7, 5.2, 1.4; H-17a), 2.06 (m; H-14a), 2.06 (m; H-15a), 1.96 (td, 12.7, 5.5; H17b), 1.87 (m; H-14b), 1.78 (dq, 14.8, 7.2; H-19a), 1.73 (dq, 14.8, 7.2; H-19b),
1.54 (td, 14.5, 4.5; H-15b), and 0.98 (t, 7.2; H3-18); 13C NMR (CD3OD, 150 MHz)
176.3 (C-21), 52.0 (C-2), 139.7 (C-13), 125.3 (C-8), 124.6 (C-11), 123.2 (C-10),
120.9 (C-9), 113.3 (C-12), 107.6 (C-7), 46.7 (C-3), 39.5 (C-20), 33.5 (C-17), 28.8
(C-15), 26.9 (C-19), 21.2 (C-16), 18.3 (C-14), and 8.3 (C-18); ESIMS (pos.) m/z
253 (M+H)+.
11. Leucomidine B (2): yellowish amorphous solid; ½a26
D 18 (c 0.3, CHCl3); IR
(KBr) mmax 3400, 1740, and 1670 cm1; UV (MeOH) kmax 298 (e 565), 222
(11250), and 202 (10800) nm; CD (MeOH) kmax 292 (De 0.57), 266 (0), 255
(+1.48), 225 (2.31), and 209 (0) nm; 1H and 13C NMR (Table 1); ESIMS (pos.) m/
z 341 (M+H)+; HRESITOFMS m/z 341.1840 (M+H)+, calcd for C20H25N2O3
341.1860.
12. Leucomidine C (3): yellowish amorphous solid; ½a26
D +32 (c 0.3, CHCl3); IR (KBr)
mmax 1740 and 1680 cm1; UV (MeOH) kmax 305 (e 1700), 273 (3250), 226
(10100), and 204 (12200) nm; 1H and 13C NMR (Table 1); ESIMS (pos.) m/z 315
(M+H)+; HRESITOFMS m/z 315.1685 (M+H)+, calcd for C18H23N2O3 315.1703.
13. Larraufie, M.-H.; Courillon, C.; Ollivier, C.; Lacôte, E.; Malacria, M.; Fensterbank,
L. J. Am. Chem. Soc. 2010, 132, 4381–4387.
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level of theory on RI-DFT B3LYP/TZVPP optimized geometries. The stable
conformation used for the DFT geometry optimization was obtained through
Monte Carlo conformational search in MacroModel 9.1 using MMFF94 force
field for the relative molecular energy evaluation.; (b) TURBOMOLE V6.3, 2011,
a development of University of Karlsruhe and Forschungszentrum Karlsruhe
GmbH, 19892007, TURBOMOLE GmbH, since 2007; available from http://
www.turbomole.com.; (c) Eickorn, K.; Treutler, O.; Ohm, H.; Haser, M.;
Ahlrichs, R. Chem. Phys. Lett. 1995, 240, 283–289; (d) Becke, A. D. Phys. Rev. A
1988, 38, 3098–3100; (e) Lee, C.; Yang, W.; Parr, R. G. Phys. Rev. B 1988, 37,
785–789; (f) Schafer, A.; Horn, H.; Ahlrichs, R. J. Chem. Phys. 1994, 100, 5829–
5835; MacroModel 9.1 (g) Mohamadi, F.; Richards, N. G. J.; Guida, W. C.;
Liskamp, R.; Lipton, M.; Caufield, C.; Chang, G.; Hendrickson, T.; Still, W. C. J.
Comput. Chem. 1990, 11, 440–467; (h) Halgren, T. J. Am. Chem. Soc. 1990, 112,
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