Synthesis of 30-amino derivatives of lupane triterpenoids

Article abstract of DOI:10.1007/s10600-006-0014-9

New derivatives of betulin and betulinic acid containing various amines on C-30 that are of interest as potentially biologically active agents were prepared. 2005 Springer Science+Business Media, Inc.

Full text of DOI:10.1007/s10600-006-0014-9

Chemistry of Natural Compounds, Vol. 41, No. 6, 2005
SYNTHESIS OF 30-AMINO DERIVATIVES OF LUPANE
TRITERPENOIDS
N. V. Uzenkova, N. I. Petrenko, M. M. Shakirov,
UDC 547.597+547.233+542.953+543.42
E. E. Shul ts, and G. A. Tolstikov
′
New derivatives of betulin and betulinic acid containing various amines on C-30 that are of interest as
potentially biologically active agents were prepared.
Key words:
betulin, betulin diacetate, methyl esters of betulinic and 3-acetylbetulinic acids, amines, NMR spectra.
Natural triterpenoids of the lupane group are of great interest because of their availability and broad spectrum of
biological activities [1-3]. Chemical transformations of these compounds at the C-3, C-20, and C-28 positions produced
synthetic derivatives that were more effective compared with the starting ones and possessed selective action [4-7]. Recent
achievements suggest that preparations based on lupane triterpenoids are promising for application to the therapy of certain
diseases [8,9].
We previouslydescribed the synthesis of amides and peptides ofbetulonic acid that contain aminoacids or their methyl
esters on C-28 [10]. It has been shown that the compounds actively inhibit reproduction of the immunodeficiency virus HIV-1
and possess other properties [11-17]. Herein we prepare derivatives containing various amines on C-30 for subsequent studies
of their pharmacological properties.
Br
30
29
CH OAc
2
CH OAc
2
Amine
NBS
AcO
AcO
1
2
30
R
R
20
29
26
21
19
17
O
18
13
22
34
11
CH OCCH
2
3
NaOH
CH OH
2
28
33
1
9
3d
15
O
3
7
27
5
CH CO
HO
3
31
32
24
4d
3a- h
23
7
5
CH Ph
a: R = N
b: R = N
c: R = N 1
d: R = N
e: R = N
O
g: R = HNCH Ph
2
2
3
1'
5
N CH
Me
1
h: R = NH
f: R = HN
3
3
3'
4'
N. N. Vorozhtsov Novosibirsk Institute of Organic Chemistry, Siberian Division, Russian Academy of Sciences,
630090, Novosibirsk, fax (3832) 30 97 52, e-mail: schultz@nioch.nsc.ru. Translated from Khimiya Prirodnykh Soedinenii,
No. 6, pp. 571-577, November-December, 2005. Original article submitted March 9, 2005.
0009-3130/05/4106-0692 ©2005 Springer Science+Business Media, Inc.
692

TABLE 1. C log P Data for Betulin and Betulinic Acid Methyl Ester and Their 30-Amino Derivatives
Compound
C log P
Compound
C log P
4g
4h
6
10
10e
10a
Betulin
4a
4b
4c
4d
8.53
8.88
9.23
10.82
6.74
7.67
9.01
9.01
8.86
8.85
7.07
8.00
9.21
4e
4f
______
10a: Methyl ester of 3β-hydroxy-30-(piperidin-1′-yl)lup-20(29)-en-28-oic acid.
10e: Methyl ester of 3β-hydroxy-30-(morpholin-1′-yl)lup-20(29)-en-28-oic acid.
TABLE 2. Physicochemical Properties of 3a-h, 4d, 9, 10
Compound
Yield, %
[α]D, °, (c)
Empirical formula
Found (M)
3a
3b
3c
3d
3e
3f
3g
3h
4d
9
63
61
52
60
61
68
45
60
93
69
95
+5 (3.38)
-7 (3.72)
+4 (2.59)
+4 (3.6)
+4 (3.72)
-6 (3.04)
-2 (2.93)
-9 (3.27)
-8 (3.6)
C39H63NO4
C40H65NO4
C46H69NO4
C39H64N2O4
C38H61NO5
C40H65NO4
C41H61NO4
C38H63NO4
C35H60N2O2
C38H62N2O4
C36H60N2O3
609.47846
623.49346
700
624.48808
611.45526
623.49326
631.45413
597.47495
540.46012
610.47111
568.45983
-5 (2.36)
-20 (3.49)
10
______
*Data for M of all compounds agreed with those calculated.
+
O
1. Diazomethane
2. NaBH
COOMe
C
COOMe
4
Ac O
2
OH
O
AcO
HO
5
6
7
Br
N
N
Me
N
N Me
COOMe
HN
N
Me
COOMe
NBS
AcO
NaOH
COOMe
HO
AcO
8
9
10
Reaction of betulin diacetate (1) with -bromosuccinimide (NBS) in CCl as before [18] produced the 30-bromo
N
4
derivative 2. Boiling 2 with various amines in alcohol gave 3a-h with secondaryor tertiaryamines on C-30. The product yields
were 45-68%. Thus, we prepared a series of compounds with an identical triterpene skeleton and various amines on C-30 of
the triterpenoid.
693

TABLE 3. Chemical Shifts of C Atoms in 13C NMR Spectra of 3a-h, 4d, 9, 10 (δ, ppm)
C atom
3a
3b
3c
3d
3e
3f
3g
3h
4d
9
10
C-1
C-2
C-3
C-4
C-5
C-6
C-7
C-8
38.26 t
23.51 t
80.76 d
37.62 s
55.23 d
18.00 t
34.04 t
40.78 s
50.16 d
36.89 s
20.76 t
26.29 t
37.38 d
42.52 s
26.95 t
29.69 t
46.12 s
49.30 d
44.52 d
38.21
23.49
80.70
37.60
55.18
17.98
33.99
40.73
50.10
36.86
20.74
26.28
37.31
42.48
26.91
29.64
46.08
49.27
44.50
38.20
23.46
80.65
37.56
55.17
17.95
33.98
40.72
50.09
36.84
20.87
26.28
37.31
42.45
26.89
29.64
46.07
49.31
44.38
151.14
31.23
34.22
27.72
16.25
15.91
15.84
14.56
62.55
109.80
63.50
-
38.17
23.43
80.62
37.53
55.14
17.92
33.95
40.69
50.05
36.81
20.67
26.31
37.26
42.43
26.85
29.60
46.05
49.32
44.31
150.67
31.23
34.19
27.69
16.24
15.90
15.81
14.57
62.48
109.89
63.32
-
38.12
23.36
80.48
37.45
55.06
17.86
33.90
40.66
50.01
36.76
20.62
26.33
37.25
42.38
26.79
29.54
46.02
49.35
44.30
150.38
31.18
34.10
27.48
16.01
15.70
15.62
14.37
62.27
109.93
63.71
-
38.17
23.48
80.65
37.58
55.18
17.98
33.98
40.73
50.09
36.85
20.69
26.27
37.32
42.47
26.87
29.58
46.10
49.21
45.06
153.00
31.17
34.26
27.75
16.29
15.97
15.86
14.57
62.42
107.14
49.80
56.09 t
38.24
23.53
80.96
37.63
55.22
17.99
34.01
40.76
50.10
36.89
20.73
26.31
37.35
42.48
26.91
29.62
46.14
49.26
44.83
152.61
31.25
34.24
27.77
16.31
15.96
15.87
14.54
62.44
107.72
52.53
38.26
23.56
80.61
37.67
55.27
18.04
34.05
40.61
50.17
36.94
20.77
26.50
37.40
42.55
26.94
29.66
46.19
49.33
44.80
153.00
31.28
34.35
27.82
16.35
16.04
15.94
14.61
62.52
107.38
48.80
38.80
27.38
78.68
38.80
55.33
18.28
33.82
40.94
50.40
37.11
20.93
26.60
37.23
42.65
27.09
29.31
47.70
49.52
44.79
150.87
31.58
34.31
27.97
15.33
16.02
16.02
14.77
59.95
109.73
63.28
-
38.21
23.48
80.71
37.59
55.22
17.97
34.07
40.49
50.27
36.89
20.80
26.21
38.06
42.17
29.55
31.92
56.40
49.78
44.22
150.78
31.76
36.68
27.74
16.29
15.99
15.77
14.61
176.42
109.88
62.80
-
38.52
27.18
78.32
38.60
55.11
18.03
34.07
40.37
50.27
36.88
20.69
26.16
37.96
42.08
29.46
31.87
56.28
49.68
44.08
150.75
31.65
36.57
27.79
15.21
15.87
15.68
14.54
176.30
109.65
62.80
-
C-9
C-10
C-11
C-12
C-13
C-14
C-15
C-16
C-17
C-18
C-19
C-20
C-21
C-22
C-23
C-24
C-25
C-26
C-27
C-28
C-29
C-30
C-1′
151.16 s 151.16
31.24 t
34.29 t
27.76 q
16.29 q
15.97 q
15.89 q
14.63 q
62.62 t
31.21
34.26
27.75
16.31
15.97
15.87
14.60
62.59
109.57 t 109.49
64.15 t
-
63.82
-
53.33 t 19.66 q
19.54 q
C-2′
C-3′
54.87 t 54.00a t 53.85a t 53.25a t 53.28a t 33.48a t
-
-
54.02 d 53.31 t
54.92 d
29.35 t
53.29
55.05
53.24
54.96
25.88 t
34.26 t
32.11 t
55.00 t
66.68 t
24.90 t
55.21 t
-
29.49 t
C-4′
C-5′
C-6′
C-7′
Ph
24.39 t
25.88 t
30.75 t
34.26 t
37.81 t
32.11 t
-
-
26.05 t
24.90 t
10.16 q
-
-
-
55.00 t
66.68 t
-
-
-
-
-
-
-
55.21 t
53.31 t
45.96 q
-
55.05
53.29
45.88
-
54.96
53.24
45.82
-
54.87 t 54.53a t 54.34a t 53.25a t 53.78a t 33.37a t
-
-
21.79 q 43.04 t 45.81 q
-
-
-
-
-
125.43
127.85
128.84
140.64
170.60
171.24
20.99
20.74
-
-
126.70
127.88
128.15
140.31
170.69
171.26
21.06
20.79
-
CH3CO at C-3 170.72 s 170.74
CH3CO at C-28 171.36 s 171.39
CH3CO at C-3 21.13 q
CH3CO at C-28 20.88 q
170.59
171.23
20.99
20.74
-
170.35
170.98
20.74
20.48
-
170.68
171.28
21.09
20.82
-
170.90
171.51
21.18
20.91
-
-
-
-
-
-
170.78
-
-
-
-
-
21.12
-
21.10
20.84
-
COOMe
-
51.04 q
51.04
______
aChemical shifts denoted with the same letters within a single column should possibly be switched.
694

The next stage of our work was to synthesize compounds containing various lupane triterpenes but the same amino
group in the 30-position in order to study the effect of the triterpene part of the aminotriterpene on the biologial activity. After
analyzing C log P of the amino derivatives of betulin and betulinic acid (Table 1), we focused on the N-methylpiperazine group.
Alkaline hydrolysis of 3d by NaOH solution (4 M) in a MeOH:THF mixture at room temperature led smoothly to
30-N′-methylpiperazino derivative 4d in 93% yield.
We also prepared N′-methylpiperazino derivatives of the methyl esters of 3-acetylbetulinic and betulinic acids. The
methyl ester of 3-acetylbetulinic acid (7) was synthesized from betulonic acid (5) in 55% yield. The N′-methylpiperazino
derivative 9 was prepared in 69% yield analogously to the amino derivative of betulin diacetate (2). Alkaline hydrolysis of 9
led smoothly to the corresponding 3-hydroxy-30-amino derivative 10 in 95% yield.
It should be noted that hydrolysis of the 30-bromo derivatives of 2 and the methyl ester of 3-acetylbetulinic acid (8)
was nonselective. In addition to the expected 30-bromo derivatives of betulin (11) and the methyl ester of betulinic acid (12),
the 30-methoxy derivatives 13 and 14, respectively, were formed. The ratio of 30-bromo derivatives 11 (12) and 30-methoxy
derivatives 13 (14) depended on the reaction conditions. Thus, after 3 d of hydrolysis at room temperature, the ratios of 11 to
13 and 12 to 14 were 1:1 and 1:2, respectively, according to PMR spectra. Increasing the hydrolysis time to 15 d gave
practically the 30-methoxy derivative. The 30-bromo derivative of the methyl ester of betulinic acid (12) could not be isolated
pure from the mixture so this compound was synthesized directly from the methyl ester of betulinic acid (6).
Br
OMe
Br
R
R
R
NaOH
+
AcO
HO
HO
2, 8
11, 12
13, 14
2: R = CH OAc
11: R = CH OH
13: R = CH OH
2
2
2
8: R = COOMe
12: R = COOMe
14: R = COOMe
The structures of all newlysynthesized compounds (2, 3a-h, 4d, 9-14) were confirmed byelemental analyses and mass
13
and NMR spectra. Signals in the C NMR spectra were assigned based on 2D spectra of 2, 3a, b, d, f, 4d, 9, 10, and 14 and
the literature [10].
EXPERIMENTAL
NMR spectra were recorded on Bruker AC-200 and AM-400 instruments at working frequencies 200.13 and
1
13
13
400.13 MHz for H and 50.32 and 100.61 MHz for C in CDCl . The multiplicity of signals in C NMR spectra was
3
determined by standard methods of recording spectra with J-modulation (JMOD) and off-resonance. Two-dimensional NMR
1
1
13
1
H— H (COSY) and C— H (COSY 125 Hz, COLOC 7 Hz) of 2, 3a, b, d, f, 4d, 9, 10, and 14 were obtained on a Bruker
1
13
DRX-500 instrument at working frequencies 500.13 MHz for H and 125.76 MHz for C in CDCl using standard Bruker
programs. The internal standards were CDCl signals (δ = 76.90) and residual H in CDCl (δ = 7.24 ppm). Mass spectra
3
1
3
C
3
H
were obtained in a Finnigan MAT 8200 high-resolution mass spectrometer with 70 eV ionzing potential. Specific rotations
([α] ) were measured on a Polamat A polarimeter in CHCl at room temperature (20-25°C). Melting points were determined
580
3
on a Kofler microheating stage. Elemental analyses were carried out in a Carlo Erba 1106 model CHN-analyzer.
The course of reactions and purityof products were monitored byTLC on Silufol UV-254 plates using solvent systems
CH Cl :MTBE (20:1) for 1, 2, and 5-8 and CHCl :CH OH (10:1) for 3a-h, 4d, and 9-14. Spots were developed by spraying
2
2
3
3
plates with H SO (20%) and heating subsequently to 100°C. Column chromatography was carried out on aluminum oxide
2
4
(activity II, CH Cl eluent).
2
2
Betulin diacetate (1) and betulonic acid (5) were prepared as before [10]. Table 2 lists the physicochemical properties
of 3a-h, 4d, 9, and 10 (yields, constants, mass spectra). Compounds 3a-h, 4d, 9, and 10 were obtained as amorphous powders.
13
Chemical shifts of characteristic protons in PMR spectra of 3c, e, g, and h are given. Table 3 lists C NMR data for 3a-h, 4d,
9, and 10. Elemental analyses of the compounds agreed with those calculated.
695

3β,28-Diacetoxy-30-bromolup-20(29)-ene (2). 1 (5.00 g, 9.50 mmol) in CCl (200 mL) was treated with NBS
4
(3.40 g, 19.10 mmol), stirred at room temperature for 5 d, and filtered through filter paper. The filtrate was evaporated. The
solid was dissolved in CH OH. The resulting solid was filtered off, washed with CH OH, and dried in a vacuum desiccator over
3
3
P O to afford 2 (3.70 g, 64%), mp 190-196°C, lit. [18] mp 185°C.
2
5
PMR spectrum (δ, ppm, J/Hz): 0.73 (1H, m, H-5), 0.77 (3H, s, Me-24), 0.78 (3H, s, Me-23), 0.79 (3H, s, Me-25), 0.91
(1H, m, H-1), 0.92 (3H, s, Me-27), 0.98 (3H, s, Me-26), 1.00 (2H, m, H-12, 15), 1.03 (1H, m, H-11), 1.10 (1H, m, H-22), 1.21
(1H, m, H-9), 1.23 (1H, m, H-16), 1.29 (1H, m, H-12), 1.32 (1H, m, H-21), 1.34 (2H, m, H-7), 1.36 (1H, m, H-11), 1.43 (2H,
m, H-6), 1.55 (2H, m, H-2), 1.60 (1H, m, H-1), 1.64 (1H, m, H-13), 1.65 (1H, m, H-15), 1.70 (1H, t, J = 11.9, H-18), 1.74 (1H,
ddd, J = 12.4, J = 8.6, J = 1.2, H-22), 1.80 (1H, ddd, J = 13.6, J = 4.7, J = 2.3, H-16), 1.98 (3H, s, AcO on C-3), 2.01 (3H,
1
2
3
1
2
3
s, AcO on C-28), 2.16 (1H, td, J = 11.2, J = 5.4, H-21), 2.39 (1H, td, J = 11.3, J = 5.4, H-19), 3.78 (1H, dd, J = 11.2,
1
2
1
2
1
J = 1.1, H -28), 3.92 (2H, AB-system, J = 10.3, H-30), 4.20 (1H, dd, J = 11.2, J = 1.8, H -28), 4.40 (1H, dd, J = 10.4,
2
A
1
2
B
1
J = 6.0, H-3), 4.97 and 5.07 (2H, both br.s, H-29).
2
13
C NMR spectrum (δ, ppm): 14.50 (q, C-27), 15.83 (q, C-26), 15.94 (q, C-25), 16.27 (q, C-24), 17.92 (t, C-6), 20.69
(t, C-11), 20.81 (q, C-34), 21.08 (q, C-32), 23.44 (t, C-2), 26.74 (t, C-12), 26.78 (t, C-15), 27.71 (q, C-23), 29.57 (t, C-16),
32.27 (t, C-21), 33.92 (t, C-7), 34.06 (t, C-22), 36.81 (s, C-10), 37.13 (t, C-30), 37.21 (d, C-13), 37.54 (s, C-4), 38.14 (t, C-1),
40.68 (s, C-8), 42.43 (s, C-14), 43.00 (d, C-19), 46.13 (s, C-17), 49.98 (d, C-9), 50.03 (d, C-18), 55.10 (d, C-6), 62.33 (t, C-28),
80.64 (d, C-3), 113.05 (t, C-29), 150.56 (s, C20), 170.75 (s, C-31), 171.30 (s, C-33). The compound was used further without
additional purification.
General Method for Preparing 30-Amino Derivatives of 1 (3a-h). 2 (1.00 g, 1.65 mmol) in alcohol (25 mL) was
treated with amine (8.25 mmol), boiled for 10 h, and poured onto ice. The resulting solid was filtered off, washed with water,
dried in a vacuum desiccator over P O , and chromatographed over Al O .
2
5
2 3
3β,28-Diacetoxy-30-(piperidin-1′-yl)lup-20(29)-ene (3a). PMR spectrum (δ, ppm, J/Hz): 0.74 (1H, m, H-5), 0.79
(3H, s, Me-24), 0.80 (6H, s, Me-23,25), 0.93 (3H, s, Me-27), 0.94 (1H, m, H-1), 0.99 (3H, s, Me-26), 1.00 (1H, m, H-15), 1.05
(1H, m, H-12), 1.10 (1H, m, H-22), 1.15 (1H, m, H-11), 1.22 (1H, m, H-16), 1.24 (1H, m, H-9), 1.29-1.42 (9H, m,
H-6,7,7,11,12,21,3′,4′,5′), 1.46 (1H, m, H-6), 1.50 (3H, m, H-3′,4′,5′), 1.57 (2H, m, H-2), 1.60 (1H, m, H-13), 1.62 (1H, m, H-1),
1.65 (1H, m, H-15), 1.69 (1H, m, H-18), 1.71 (1H, m, H-22), 1.80 (1H, m, H-16), 1.99 (3H, s, AcO on C-3), 2.01 (1H, m, H-21),
2.02 (3H, s, AcO on C-28), 2.32 (5H, br.s, H-19,2′,2′,6′,6′), 2.75 (2H, AB-system, J = 14.0, H-30), 3.80 (1H, dd, J = 11.0,
1
J = 1.9, H -28), 4.21 (1H, dd, J = 11.0, J = 1.3, H -28), 4.42 (1H, dd, J = 10.3, J = 5.8, H-3), 4.80 and 4.81 (2H, both br.s,
2
A
1
2
B
1
2
H-29).
3β,28-Diacetoxy-30-(4′-methylpiperidin-1′-yl)lup-20(29)-ene (3b). PMR spectrum (δ, ppm, J/Hz): 0.73 (1H, m,
H-5), 0.79 (9H, s, Me-23,24,25), 0.87 (3H, d, J = 6.5, Me-7′), 0.91 (1H, m, H-1), 0.92 (3H, s, Me-27), 0.98 (3H, s, Me-26), 1.00
(1H, m, H-15), 1.04 (1H, m, H-12), 1.08 (1H, m, H-22), 1.15 (2H, m, H-3′,5′), 1.16 (1H, m, H-11), 1.21 (1H, m, H-16), 1.23
(2H, m, H-9,4′), 1.26-1.41 (6H, H-6,7,7,11,12,21), 1.45 (1H, m, H-6), 1.51 (2H, m, H-3′,5′), 1.55 (2H, m, H-2), 1.58 (1H, m,
H-13), 1.61 (1H, m, H-1), 1.64 (1H, m, H-15), 1.68 (1H, m, H-18), 1.70 (1H, m, H-22), 1.77 (1H, m, H-2′ or 6′), 1.79 (1H, m,
H-16), 1.85 (1H, m, H-2′ or 6′), 1.99 (3H, s, AcO on C-3), 2.00 (1H, m, H-21), 2.02 (3H, s, AcO on C-28), 2.33 (1H, m, H-19),
2.79 (4H, m, H-30,30,2′,6′), 3.79 (1H, d, J = 11.0, H -28), 4.20 (1H, d, J = 11.4, H -28), 4.42 (1H, dd, J = 10.4, J = 6.0, H-3),
A
B
1
2
4.80 (2H, br.s, H-29).
3β,28-Diacetoxy-30-(4′-benzylpiperidin-1′-yl)lup-20(29)-ene (3c). PMRspectrum (δ, ppm, J/Hz): 0.76(1H, m, H-5),
0.81 (9H, s, Me-23,24,25), 0.93 (3H, s, Me-27), 1.01 (3H, s, Me-26), 2.00 (3H, s, AcO on C-3), 2.03 (3H, s, AcO on C-28), 2.34
(1H, m, H-19), 2.50 (2H, m, CH ), 2.79 (4H, m, H-30, CH Ph), 3.81 (1H, d, J = 11.0, H -28), 4.20 (1H, d, J = 11.0, H -28),
2
2
A
B
4.44 (1H, dd, J = 10.5, J = 6.1, H-3), 4.84 (2H, br.s, H-29), 7.20 (5H, m, Ph).
1
2
3β,28-Diacetoxy-30-(4 -methylpiperazin-1 -yl)lup-20(29)-ene (3d). PMR spectrum (δ, ppm, J/Hz): 0.70 (1H, m,
H-5), 0.75 (3H, s, Me-24), 0.76 (3H, s, Me-23), 0.77 (3H, s, Me-25), 0.89 (3H, s, Me-27), 0.95 (3H, s, Me-26), 0.90 (1H, m,
H-1), 0.97 (1H, m, H-15), 1.01 (1H, m, H-12), 1.05 (1H, m, H-22), 1.12 (1H, m, H-11), 1.18 (1H, m, H-16), 1.20 (1H, m, H-9),
1.24-1.37 (6H, m, H-6,7,7,11,12,21), 1.43 (1H, m, H-6), 1.54 (2H, m, H-2), 1.56 (1H, m, H-13), 1.58 (1H, m, H-1), 1.60 (1H,
m, H-15), 1.64 (1H, m, H-18), 1.68 (1H, m, H-22), 1.77 (1H, m, H-16), 1.96 (3H, s, AcO on C-3), 1.97 (1H, m, H-21), 1.99 (3H,
s, AcO on C-28), 2.20 (3H, s, N-Me), 2.29 (1H, m, H-19), 2.36 (8H, m, H-2′,3′,5′,6′), 2.76 (2H, AB-system, J = 14.0, H-30),
3.76 (1H, d, J = 11.2, H -28), 4.17 (1H, d, J = 11.1, H -28), 4.39 (1H, dd, J = 10.5, J = 5.8, H-3), 4.78 and 4.80 (2H, both
A
B
1
2
br.s, H-29).
696

3β,28-Diacetoxy-30-(morpholin-1′-yl)lup-20(29)-ene (3e). PMR spectrum (δ, ppm, J/Hz): 0.72 (1H, m, H-5), 0.76
(6H, s, Me-23,24), 0.78 (3H, s, Me-25), 0.91 (3H, s, Me-27), 0.96 (3H, s, Me-26), 1.93 (3H, s, AcO on C-3), 1.96 (3H, s, AcO
on C-28), 2.32 (5H, m, H-19,2′,6′), 2.76 (2H, m, H-30), 3.57 (4H, m, H-3′,5′), 3.78 (1H, d, J = 10.6, H -28), 4.18 (1H, d,
A
J = 10.6, H -28), 4.46 (1H, dd, J = 10.8, J = 6.0, H-3), 4.79 and 4.81 (2H, both br.s, H-29).
B
1
2
3β,28-Diacetoxy-30-cyclohexylaminolup-20(29)-ene (3f). PMR spectrum (δ, ppm, J/Hz): 0.73 (1H, m, H-5), 0.77
(3H, s, Me-24), 0.78 (3H, s, Me-23), 0.79 (3H, s, Me-25), 0.90 (1H, m, H-1), 0.91 (3H, s, Me-27), 0.97 (3H, s, Me-26), 0.99
(1H, m, H-15), 1.04 (1H, m, H-12), 1.08 (1H, m, H-22), 1.12 (4H, m, H-2′,3′,5′,6′), 1.16 (2H, m, H-11,4′), 1.20 (1H, m, H-16),
1.23 (1H, m, H-9), 1.28-1.40 (6H, m, H-6,7,7,11,12,21), 1.45 (1H, m, H-6), 1.56 (2H, m, H-2), 1.57 (2H, m, H-13,4′), 1.60 (1H,
m, H-1), 1.64 (1H, m, H-15), 1.67 (2H, m, H-3′,5′), 1.69 (1H, m, H-18), 1.71 (1H, m, H-22), 1.79 (1H, m, H-16), 1.82 (2H, m,
H-2′,6′), 1.98 (3H, s, AcO on C-3), 2.00 (1H, m, H-21), 2.01 (3H, s, AcO on C-28), 2.28 (1H, m, H-19), 2.37 (1H, m, H-1′),
3.15 (2H, AB-system, J = 15.3, H-30), 3.78 (1H, d, J = 11.0, H -28), 4.18 (1H, d, J = 11.1, H -28), 4.40 (1H, dd, J = 10.6,
A
B
1
J = 5.8, H-3), 4.76 and 4.79 (2H, both br.s, H-29).
2
3β,28-Diacetoxy-30-benzylaminolup-20(29)-ene (3g). PMR spectrum (δ, ppm, J/Hz): 0.70 (1H, m, H-5), 0.82 (9H,
s, Me-23,24,25), 0.90 (3H, s, Me-27), 1.00 (3H, s, Me-26), 2.00 (3H, s, AcO on C-3), 2.03 (3H, s, AcO on C-28), 2.30 (1H, m,
H-19), 3.18 (2H, m, CH Ph), 3.80 (3H, m, H-28,30), 4.22 (1H, d, J = 10.8, H-28), 4.44 (1H, dd, J = 10.8, J = 6.0, H-3), 4.85
2
1
2
and 4.89 (2H, both br.s, H-29), 7.25 (5H, m, Ph).
3β,28-Diacetoxy-30-(sec-butyl)aminolup-20(29)-ene (3h). PMR spectrum (δ, ppm, J/Hz): 0.75 (1H, m, H-5), 0.81
(9H, s, Me-23,24,25), 0.88 (3H, t, J = 7.6, Me-4′), 0.94 (3H, s, Me-27), 1.00 (3H, s, Me-26), 1.04 (3H, d, J = 6.1, Me-1′), 2.04
(3H, s, AcO on C-3), 2.05 (3H, s, AcO on C-28), 2.32 (1H, m, H-19), 2.57 (1H, m, H-2′), 3.16 (2H, m, H-30), 3.80 (1H, d,
J = 10.6, H -28), 4.23 (1H, d, J = 10.6, H -28), 4.43 (1H, dd, J = 10.8, J = 5.8, H-3), 4.83 (2H, br.s, H-29).
A
B
1
2
3β,28-Hydroxy-30-(4′-methylpiperazin-1′-yl)lup-20(29)-ene (4d). 3d(0.20g, 0.32mmol)in MeOH(4mL)andTHF
(2 mL) under Ar at 0°C was treated with NaOH solution (0.32 mL, 1.32 mmol, 4 N), held at room temperature for 2 d, and
poured onto ice. The resulting solid was filtered off, washed with water, and dried in air.
PMR spectrum (δ, ppm, J/Hz): 0.61 (1H, m, H-5), 0.70 (3H, s, Me-24), 0.76 (3H, s, Me-25), 0.90 (3H, s, Me-23), 0.92
(3H, s, Me-27), 0.96 (3H, s, Me-26), 0.97 (1H, m, H-15), 0.99 (1H, m, H-22), 1.02 (1H, m, H-12), 1.13 (1H, m, H-16), 1.15 (1H,
m, H-11), 1.20 (1H, m, H-9), 1.25-1.40 (6H, m, H-6,7,7,11,12,21), 1.46 (1H, m, H-6), 1.52 (2H, m, H-2), 1.54 (1H, m, H-13),
1.57 (1H, m, H-1), 1.60 (1H, m, H-18), 1.64 (1H, m, H-15), 1.80 (1H, m, H-22), 1.89 (1H, m, H-16), 1.99 (1H, m, H-21), 2.20
(3H, s, N-Me), 2.23 (1H, m, H-19), 2.38 (8H, br.s, H-2′,3′,5′,6′), 2.79 (2H, AB-system, J = 14.2, H-30), 3.10 (1H, dd, J = 11.0,
1
J = 5.4, H-3), 3.23 (1H, d, J = 10.7, H -28), 3.69 (1H, d, J = 10.7, H -28), 4.79 and 4.81 (2H, both br.s, H-29).
2
A
B
Methyl Ester of 3β-Acetoxylup-20(29)-en-28-oic Acid (7). 5 (3.88 g, 8.55 mmol) was dissolved in anhydrous
diethylether (235 mL), cooled to 0°C, stirred vigorously, treated dropwise with diazomethane until a yellow color was stable,
held at 0°C for 48 h, dried over anhydrous MgSO , and evaporated. The solid was washed with MeOH and chromatographed
4
over Al O (CH Cl eluent). The purified methyl ester of betulonic acid (2.81 g, 6.00 mmol) was dissoslved in dry THF
2
3
2
2
(140 mL), cooled to 0°C, treated in portions with NaBH (0.98 g, 25.79 mmol), stirred with cooling for 3 h, held at room
4
temperature for 1 d, and poured onto ice with dilute HCl. The resulting solid was filtered off, washed with water, dried in a
desiccator over P O , and chromatographed over Al O . A mixture of purified methyl ester of betulinic acid (6, 2.30 g,
2
5
2 3
4.90 mmol), Ac O (18 mL), and pyridine (70 mL) was held at room temperature, periodically stirred during 3 d, and poured
2
onto ice with conc. HCl. The resulting solid was filtered off, washed with water, and dried over P O to afford the methyl ester
2
5
of 3-acetylbetulinic acid (7, 2.40 g, 55%), mp 201-203°C, lit. [19] mp 202-203°C.
Methyl Ester of 3β-Acetoxy-30-bromolup-20(29)-en-28-oic Acid (8). A mixture of 7 (2.40 g, 4.69 mmol) and NBS
(1.67 g, 9.38 mmol) in CCl (96 mL) was held at room temperature, periodically stirred during 3 d, and filtered through filter
4
paper. The filtrate was evaporated. The solid was dissolved in MeOH. The resulting solid was filtered off, washed with MeOH,
and dried in a vacuum desiccator over P O to afford 8 (1.47 g, 53%), mp 218-220°C, [α]D -5° (c 2.93), lit. [20] mp 235-236°C,
2
5
[α]D +42.55. Found, %: Br 13.60; M-Br (mass spectrometry) 511.38024. C H O Br. Calc., %: Br 13.51; M-Br 511.37871.
33 51
4
PMR spectrum (δ, ppm, J/Hz): 0.72 (1H, m, H-5), 0.77 (3H, s, Me-24), 0.78 (6H, s, Me-23,25), 0.85 (3H, s, Me-26),
0.91 (3H, s, Me-27), 1.97 (3H, s, AcO), 2.98 (1H, td, J = 11.7, J = 4.6, H-19), 3.61 (3H, s, OMe), 3.93 (2H, br.s, H-30), 4.40
1
2
(1H, dd, J = 10.5, J = 5.8, H-3), 4.98 and 5.08 (2H, both br.s, H-29).
1
13
2
C NMR spectrum (δ, ppm): 14.49 (q, C-27), 15.79 (q, C-26), 15.98 (q, C-25), 16.30 (q, C-24), 18.00 (t, C-6), 20.84
(t, C-11), 21.06 (q, C-32), 23.50 (t, C-2), 26.63 (t, C-12), 27.76 (q, C-23), 29.49 (t, C-15), 31.84 (t, C-16), 32.91 (t, C-21), 34.13
(t, C-7), 36.45 (t, C-22), 36.74 (t, C-30), 36.93 (s, C-10), 37.60 (s, C-4), 38.07 (d, C-13), 38.24 (t, C-1), 40.54 (s, C-8), 42.17
697

(s, C-14), 42.97 (d, C-19), 50.26 (d, C-9), 50.86 (d, C-18), 51.10 (q, OMe), 55.26 (d, C-5), 56.37 (s, C-17), 80.67 (d, C-3),
113.20 (t, C-29), 151.14 (s, C-20), 170.65 (s, C-31), 176.13 (s, C-28). The compound was used further without additional
purification.
Methyl Ester of 3β-Acetoxy-30-(4′-methylpiperazin-1′-yl)lup-20(29)-en-28-oic Acid (9). A solution of 8 (0.90 g,
1.52 mmol) in alcohol (23 mL) was treated with 1-methylpiperazine (0.84 mL, 7.59 mmol), boiled for 10 h, and poured onto
ice. The resulting solid was filtered off, washed with water, dried in a desiccator over P O , and chromatographed over Al O .
2
5
2 3
PMR spectrum (δ, ppm, J/Hz): 0.72 (1H, m, H-5), 0.76 (3H, s, Me-24), 0.78 (6H, s, Me-23,25), 0.84 (3H, s, Me-26),
0.90 (3H, s, Me-27), 0.92 (1H, m, H-1), 0.98 (1H, m, H-12), 1.08 (1H, m, H-15), 1.17 (1H, m, H-11), 1.22 (1H, m, H-9), 1.25-
1.36 (7H, m, H-6,7,7,11,15,16,21), 1.38 (1H, m, H-22), 1.40-1.47 (2H, m, H-6,12), 1.55 (2H, m, H-2), 1.61 (1H, m, H-1), 1.66
(1H, t, J = 11.3, H-18), 1.77 (1H, dd, J = 12.3, J = 7.7, H-22), 1.88 (1H, m, H-21), 1.97 (3H, s, AcO), 2.10 (1H, td, J = 12.1,
1
2
1
J = 3.6, H-13), 2.18 (1H, m, H-16), 2.23 (3H, s, N-Me), 2.37 (8H, br.s, H-2′,3′,5′,6′), 2.80 (2H, AB-system, J = 14.3, H-30),
2
2.86 (1H, td, J = 11.0, J = 4.4, H-19), 3.60 (3H, s, OMe), 4.40 (1H, dd, J = 10.3, J = 6.1, H-3), 4.80 and 4.84 (2H, both br.s,
1
2
1
2
H-29).
Methyl Ester of 3β-Hydroxy-30-(4′-methylpiperazin-1′-yl)lup-20(29)-en-28-oic Acid (10). A solution of 9
(0.22 g, 0.36 mmol) in MeOH (4.4 mL) and THF (2.2 mL) under Ar at 0 C was treated with NaOH solution (0.36 mL,
1.44 mmol, 4 N), held at room temperature for 2 d, and poured onto ice. The resulting solid was filtered off, washed with water,
and dried in air.
PMR spectrum (δ, ppm, J/Hz): 0.57 (1H, m, H-5), 0.65 (3H, s, Me-24), 0.71 (3H, s, Me-25), 0.79 (1H, m, H-1), 0.80
(3H, s, Me-26), 0.86 (6H, s, Me-23,27), 0.94 (1H, m, H-12), 1.05 (1H, m, H-15), 1.14 (2H, m, H-9,11), 1.20-1.33 (7H, m,
H-6,7,7,11,15,16,21), 1.33-1.45 (3H, m, H-6,12,22), 1.48 (2H, m, H-2), 1.56 (1H, m, H-1), 1.61 (1H, t, J = 11.4, H-18), 1.73
(1H, m, H-22), 1.84 (1H, m, H-21), 2.06 (1H, td, J = 12.0, J = 2.6, H-13), 2.14 (1H, m, H-16), 2.18 (3H, s, N-Me), 2.35 (8H,
1
2
br.s, H-2′,3′,5′,6′), 2.77 (2H, AB-system, J = 14.3, H-30), 2.82 (1H, td, J = 11.2, J = 4.0, H-19), 3.05 (1H, dd, J = 10.3,
1
2
1
J = 5.8, H-3), 3.56 (3H, s, OMe), 4.76 and 4.80 (2H, both br.s, H-29).
2
General Method for Hydrolysis of 30-Bromo Derivatives (2, 8). A solution of 2 (8) (1.9 mmol) in MeOH (26 mL)
and THF (12 mL) under Ar at 0°C was treated with NaOH solution (1.2 mL, 4.8 mmol, 4 N), held at room temperature for
3 d, and poured onto ice with dilute HCl. The resulting solid was filtered off, washed with water, and dried over P O to afford
2
5
a mixture of 11 and 13 or 12 and 14 in ~1:1 or 1:2 ratios, respectively, according to PMR spectra. Chromatography over Al O
2
3
produced analytically pure compounds.
3β,28-Hydroxy-30-bromolup-20(29)-ene (11). Mp 201-204°C, [α]D -2° (c 3.49). Mass spectrum, m/z: 520.29120
+
+
[M] . C H O Br. Calc.: 520.29162 [M] .
30 49
2
PMR spectrum (δ, ppm, J/Hz): 0.66 (1H, m, H-5), 0.73 (3H, s, Me-24), 0.80 (3H, s, Me-25), 0.94 (3H, s, Me-23), 0.96
(3H, s, Me-27), 1.08 (3H, s, Me-26), 3.14 (1H, dd, J = 10.1, J = 5.2, H-3), 3.29 (1H, d, J = 10.8, H -28), 3.76 (1H, d, J = 10.9,
1
2
A
H -28), 3.99 (1H, m, H-30), 5.00 and 5.09 (2H, both br.s, H-29).
B
13
C NMR spectrum (δ, ppm): 14.64 (q, C-27), 15.24 (q, C-24), 15.95 (q, C-25,26), 18.30 (t, C-6), 20.85 (t, C-11),
26.92 (t, C-12,15), 27.25 (t, C-2), 27.88 (q, C-23), 29.15 (t, C-16), 32.54 (t, C-21), 33.59 (t, C-22), 34.19 (t, C-7), 37.04 (t,
C-30), 37.11 (s, C-10), 37.20 (d, C-13), 38.62 (t, C-1), 38.73 (s, C-4), 40.86 (s, C-8), 42.59 (s, C-14), 43.38 (d, C-19), 47.70
(s, C-17), 50.22 (d, C-9,18), 55.19 (d, C-5), 60.13 (t, C-28), 78.81 (d, C-3), 113.12 (t, C-29), 150.92 (s, C-20).
3β,28-Hydroxy-30-methoxylup-20(29)-ene (13). Mp 212-214°C, [α]D -13° (c 3.38). Mass spectrum, m/z: 472.40779
+
+
[M] . C H O . Calc.: 472.39162 [M] .
31 52
3
PMR spectrum (δ, ppm, J/Hz): 0.66 (1H, m, H-5), 0.74 (3H, s, Me-24), 0.80 (3H, s, Me-25), 0.94 (3H, s, Me-23), 0.96
(3H, s, Me-27), 1.00 (3H, s, Me-26), 3.16 (1H, dd, J = 10.3, J = 5.1, H-3), 3.29 (1H, d, J = 10.8, H -28), 3.33 (3H, s, OMe),
1
2
A
3.76 (1H, d, J = 10.9, H -28), 3.84 (1H, m, H-30), 4.88 and 4.90 (2H, both br.s, H-29).
B
13
C NMR spectrum (δ, ppm): 14.62 (q, C-27), 15.23 (q, C-24), 15.96 (q, C-25,26), 18.20 (t, C-6), 20.86 (t, C-11),
26.62 (t, C-12), 26.98 (t, C-15), 27.29 (t, C-2), 27.88 (q, C-23), 29.18 (t, C-16), 31.44 (t, C-21), 33.70 (t, C-22), 34.22 (t, C-7),
37.07 (s, C-10), 37.15 (d, C-13), 38.64 (t, C-1), 38.74 (s, C-4), 40.88 (s, C-8), 42.59 (s, C-14), 43.61 (d, C-19), 47.67 (s, C-17),
49.37 (d, C-18), 50.31 (d, C-9), 55.21 (d, C-5), 58.12 (q, OMe), 60.14 (t, C-28), 74.86 (t, C-30), 78.81 (d, C-3), 108.99 (t,
C-29), 150.97 (s, C-20).
Methyl Ester of 3β-Hydroxy-30-methoxylup-20(29)-en-28-oic Acid (14). Amorphous powder, [α]D -20° (c 8.11).
+
+
Mass spectrum, m/z: 500.39376 [M] . C H O . Calc.: 500.38654 [M] .
32 52
4
698

PMR spectrum (δ, ppm, J/Hz): 0.62 (1H, m, H-5), 0.71 (3H, s, Me-24), 0.77 (3H, s, Me-25), 0.85 (1H, td, J = 12.4,
1
J = 4.6, H-1), 0.86 (3H, s, Me-26), 0.91 (3H, s, Me-23), 0.92 (3H, s, Me-27), 1.05 (1H, m, H-15), 1.10 (1H, m, H-12), 1.20 (2H,
2
m, H-9,11), 1.25-1.40 (6H, m, H-7,7,11,15,16,21), 1.40-1.53 (4H, m, H-6,6,12,22), 1.55 (2H, m, H-2) 1.61 (1H, td, J = 13.0,
1
J = 3.6, H-1), 1.65 (1H, t, J = 11.4, H-18), 1.81 (1H, dd, J = 11.9, J = 7.9, H-22), 1.92 (1H, m, H-21), 2.12 (1H, td, J = 12.8,
2
1
2
1
J = 3.6, H-13), 2.20 (1H, dd, J = 8.9, J = 2.7, H-16), 2.83 (1H, td, J = 11.3, J = 4.5, H-19), 3.12 (1H, dd, J = 11.3,
2
1
2
1
2
1
J = 5.0, H-3), 3.30 (3H, s, OMe), 3.61 (3H, s, COOMe), 3.82 (1H, br.s, H-30), 4.88 and 4.89 (2H, both br.s, H-29).
2
13
C NMR spectrum (δ, ppm): 14.53 (q, C-27), 15.21 (q, C-24), 15.81 (q, C-26), 15.95 (q, C-25), 18.14 (t, C-6), 20.83
(t, C-11), 26.48 (t, C-12), 27.25 (t, C-2), 27.84 (q, C-23), 29.56 (t, C-15), 31.94 (t, C-16,21), 34.22 (t, C-7), 36.62 (t, C-22),
37.04 (s, C-10), 38.11 (d, C-13), 38.59 (t, C-1), 38.69 (s, C-4), 40.55 (s, C-8), 42.20 (s, C-14), 42.94 (d, C-19), 49.92 (d, C-18),
50.40 (d, C-9), 51.08 (q, OMe), 55.23 (d, C-5), 56.40 (s, C-17), 58.14 (q, COOMe), 74.86 (t, C-30), 78.73 (d, C-3), 108.61 (t,
C-29), 151.13 (s, C-20), 176.37 (s, C-28).
Methyl Ester of 3β-Hydroxy-30-bromolup-20(29)-en-28-oic Acid (12). A mixture of 6 (0.55 g, 1 mmol) and NBS
(0.36 g, 2 mmol) in CCl (22 mL) was held at room temperature, periodically stirred during 1.5 d, and filtered through filter
4
paper. The filtrate was evaporated. The solid was chromatographed over Al O to afford 12 (0.25 g, 39%) as an amorphous
2
3
powder, [α]D -12° (c 4.05).
PMR spectrum (δ, ppm, J/Hz): 0.64 (1H, m, H-5), 0.72 (3H, s, Me-24), 0.78 (3H, s, Me-25), 0.87 (3H, s, Me-26), 0.92
(3H, s, Me-23), 0.94 (3H, s, Me-27), 3.00 (1H, td, J = 11.0, J = 4.8, H-19), 3.14 (1H, dd, J = 11.0, J = 5.2, H-3), 3.64 (3H,
1
2
1
2
s, OMe), 3.95 (2H, s, H-30), 5.00 and 5.10 (2H, both br.s, H-29).
13
C NMR spectrum (δ, ppm): 14.58 (q, C-27), 15.21 (q, C-24), 15.84 (q, C-26), 15.95 (q, C-25), 18.16 (t, C-6), 20.87
(t, C-11), 26.71 (t, C-12), 27.25 (t, C-2), 27.86 (q, C-23), 29.55 (t, C-15), 31.92 (t, C-16), 32.92 (t, C-21), 34.26 (t, C-7), 36.52
(t, C-22), 36.67 (t, C-30), 37.08 (s, C-10), 38.17 (d, C-13), 38.63 (t, C-1), 38.71 (s, C-4), 40.59 (s, C-8), 42.24 (s, C-14), 43.11
(d, C-19), 50.42 (d, C-9), 50.92 (d, C-18), 51.15 (q, COOMe), 55.26 (d, C-5), 56.44 (s, C-17), 78.82 (d, C-3), 113.26 (t, C-29),
151.20 (s, C-20), 176.22 (s, C-28).
ACKNOWLEDGMENT
The work was supported financially by the Russian Foundation for Basic Research (grant No. 03-33093) and a grant
to leading science schools NSh-1488.2003.3.
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