Triterpene glycosides from Tragacantha stipulosa and their genins. Structure of cyclostipuloside E

Article abstract of DOI:10.1023/B:CONC.0000025463.46079.8f

The known compound trojanoside A(1) and a new cycloartane glycoside cyclostipuloside E (2) were isolated from the aerial parts of Tragacantha stipulosa Boriss. The structure of cyclostipuloside E was proposed as 24R-cycloartan-3β,6α,16β,24,25-pentaol6,16,25-tri-O-β-D- glucopyranoside3-O-β-D-xylopyranoside based on physicochemical data and chemical transformations.

Full text of DOI:10.1023/B:CONC.0000025463.46079.8f

Chemistry of Natural Compounds, Vol. 40, No. 1, 2004
TRITERPENE GLYCOSIDES FROM Tragacantha stipulosa AND
THEIR GENINS. STRUCTURE OF CYCLOSTIPULOSIDE E
T. N. Kaipnazarov, K. K. Uteniyazov, and Z. Saatov
UDC 547.918:547.926
1
2
The known compound trojanoside A ( ) and a newcycloartane glycoside cyclostipuloside E ( ) were isolated
Tragacantha stipulosa
from the aerial parts of
Boriss. The structure of cyclostipuloside E was proposed as
β α
β
β
β
24R-cycloartan-3 ,6 ,16 ,24,25-pentaol 6,16,25-tri-O- -D-glucopyranoside 3-O- -D-xylopyranoside based
on physicochemical data and chemical transformations.
Key words: cycloartanes, trojanoside A, cyclosiversioside F, cyclosiversigenin, cyclostipuloside E, cycloasgenin C,
cycloasgenin C monoside.
In continuation of research on cycloartane triterpenoids from
Boriss (Leguminosae) [1], we
Tragacantha stipulosa
isolated the known cycloartane glycoside trojanoside A (1) and a new compound, cyclostipuloside E (2), from the butanol
fraction of the methanol extract of the aerial parts. Six compounds were previouslyisolated from this plant: cycloglobiseposide
B [2], cyclostipuloside A, askendoside G, askendoside D [3], cyclostipuloside D [4], and cyclostipuloside C [1].
The PMR spectrum of 1 [5] contains signals for seven methyls at 0.89, 0.94, 0.98, 1.02, 1.19, 1.24, and 1.24 ppm in
addition to signals for two 1H doublets of an AB system at 0.17 and 0.47 ppm, which are unambiguouslyassigned to methylene
H atoms of the cyclopropane ring. Therefore, 1 is a cycloartane.
The PMR spectrum of this compound contains a 3H singlet at 1.95 ppm that indicates the molecule has a single acetyl.
13
This is also evident in the C NMR spectrum, in which resonances of C atoms of a single acetate are clearly visible at 21.24
-1
and 171.25 ppm. This is confirmed by the presence in the IR spectrum of an absorption band for an ester at 1717 cm .
Saponification of 1 by base produced the deacetylated derivative 3, which coincided on TLC with cyclosiversioside F
[6]. Acid hydrolysis showed that the genin is cyclosiversigenin (4) whereas the carbohydrate consists of xylose and glucose.
OH
OH
OH
H
H
H
O
O
O
OAc
OH
OH
+
H
KOH
HO
p
β
p β
Xyl -D- -O
Xyl -D- -O
OH
O-β-D-Glcp
O-β-D-Glcp
1
3
4
13
Comparison of the C NMR spectra of 1 and 3 revealed that all signals in general coincide. The only difference is
that the signal for C-16 in the spectrum of 1 is shifted to weak field by +1.54 ppm.
It can be concluded from the above that the acetyl in 1 is located on C-16 of the aglycon.
S. Yu. Yunusov Institute of the Chemistry of Plant Substances, Academy of Sciences of the Republic of Uzbekistan,
700170, Tashkent, fax (99871) 120 64 75, e-mail: root@icps.org.uz. Translated from Khimiya Prirodnykh Soedinenii, No. 1,
pp. 35-38, January-February, 2004. Original article submitted November 3, 2003.
0009-3130/04/4001-0040 ©2004 Plenum Publishing Corporation
40

TABLE 1. 13C NMR Chemical Shifts (ppm) of Cyclostipuloside E (2), Cycloasgenin C (5), and its Monoside
(6) in C5D5N
Compound
C atom
2
5
6
1
2
3
4
5
6
7
8
32.57
30.05
88.77
42.72
54.04
79.32
38.39
46.86
21.28
29.60
26.26
32.83
45.63
46.86
47.30
82.84
57.51
18.21
30.37
31.74
19.04
30.22
34.55
78.86
80.79
22.32
23.50
20.23
28.92
16.74
32.81
31.48
78.84
42.43
53.88
68.25
38.56
47.02
21.07
29.55
26.26
34.37
45.55
46.90
47.82
71.77
57.19
18.08
30.30
31.96
19.18
30.63
34.37
79.96
72.70
25.42
26.39
20.25
29.44
16.23
32.61
30.10
88.75
42.71
54.10
67.89
38.40
46.92
21.15
29.62
26.25
32.80
45.61
46.92
47.39
71.75
57.15
18.52
30.25
31.95
19.14
30.49
34.57
78.82
72.65
24.83
25.18
20.24
29.04
16.65
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
3-O-β-D-Xylp
6-O-β-D-Glcp
1′
2′
3′
4′
5′
107.60
75.62
78.04
71.85
67.05
107.59
75.63
78.44
71.33
67.05
1″
2″
3″
4″
5″
6″
105.68
75.85
77.62
72.27
78.79
62.27
41

TABLE 1. (Continued)
C atom
Compound
2
5
6
16-O-β-D-Glcp
1′″
2′″
3′″
4′″
5′″
6′″
106.28
75.94
78.49
71.27
78.14
62.94
25-O-β-D-Glcp
1″″
2″″
3″″
4″″
5″″
6″″
98.75
75.35
78.72
71.85
78.32
62.87
13
The structure of 1 was confirmed using PMR, C NMR, and two-dimensional spectra (COSY, TOCSY). Comparison
of the C NMR spectrum of the genin of compound 1 with those in the literature established that it is identical to that of
cyclosiversigenin (4) [6, 7].
13
Thus, the structure of trojanoside A (1) is 16-O-acetyl-20 ,24 -epoxycycloartan-3β,6α,16β,25-tetraol 3-O-β-D-
R
S
xylopyranoside 6-O-β-D-glucopyranoside.
The PMR spectrum of the new compound, cyclostipuloside E (2), hasat strong field signals for protons of seven methyls
2
in addition to 1H doublets for an AB system at 0.25 and 0.45 ppm with J = 3.9 Hz, which are characteristic of methylene
-1
protons of a cyclopropane ring. The presence of a three-membered ring was confirmed by an absorption band at 2972 cm in
the IR spectrum of 2.
Acid hydrolysis of 2 produced the genin, which was identified using spectral and literature data as cycloasgenin C (5)
[2]. Paper chromatography (PC) of the hydrolysates detected D-xylose and D-glucose.
13
The PMR and C NMR spectra of 2 exhibit signals for four anomeric protons at 4.78, 4.84, 4.92, and 5.20 ppm and
four anomeric C atoms that resonate at 107.60, 105.68, 106.28, and 98.75 ppm, respectively(Table 1). These data indicate that
2 is a tetraoside.
13
Comparison of the C NMR spectra of 5 [2] and 2 showed that four C atoms of the genin (C-3, C-6, C-16, and C-25)
are affected by glycosylation and resonate at 88.77, 79.32, 82.84, and 80.79 ppm, respectively.
Thus, it can be assumed that the sugars are bonded to the genin through hydroxyls on C-3, C-6, C-16, and C-25.
Enzymatic hydrolysis of 2 by gastric juice of the snail
produced progenin 6 [2].
Helix plectotropis
Acid hydrolysis of 6 gave cycloasgenin C (5) [2]. PC of the hydrolysate detected D-xylose.
2
The PMR spectrum of 6 [2] exhibits a signal for an anomeric proton as a doublet at 4.86 ppm with SSCC J = 7.7 Hz.
13
Comparison of the C NMR spectra of 6 [2] and 5 [2] found that the signal for C-3 undergoes a paramagnetic shift by9.93 ppm
compared with the genin (Table 1).
Therefore, 6 [2] contains D-xylose bonded to the genin through the C-3 hydroxyl and is cycloasgenin C 3-O-β-D-
xylopyranoside [2].
These data indicate that the three glucoses in 2 are bonded to hydroxyls on C-6, C-16, and C-25. This conclusion was
13
4
confirmed using PMR and C NMR spectra of 2. The SSCC of the anomeric protons are consistent with the C -conformation
1
of the pyranose rings and the -configuration of the glycoside centers.
Thus, 2 has the structure 24R-cycloartan-3β,6α,16β,24,25-pentaol 6,16,25-tri-O-β-D-glucopyranoside 3-O-β-D-
xylopyranoside.
42

OH
OH
O
D Glcp
OH
β
O
OH
D Glcp
β
Enzyme
O
Xyl
D
β
p
Xylp-D-β -O
OH
O-β-D-Glcp
6
2
+
OH
OH
H
+
H
OH
HO
OH
5
EXPERIMENTAL
For general comments, see [1].
Separation of the Butanol Fraction. The butanol fraction was chromatographed over a column using
CHCl :CH OH:H O (70:23:3) to afford trojanoside A (1, 1.5 g, 0.06%) [5] and 2 (90 mg, 0.0036%) (here and hereinafter yields
3
3
2
are calculated per air-dried raw material).
25
Trojanoside A (1). C H O , mp 292-285°C (CH OH), [α]
+20.1° ( 0.1, CH OH). IR spectrum (KBr,
c
3
43 70 15
3
D
ν
, cm-1): 3416 (OH), 3030 (cyclopropane), 1717 (C=O), 1272 and 1043 (C–O–C).
max
PMR spectrum (DMSO, δ, ppm, 0 = TMS): 1.44, 1.16 (H-1), 1.82, 1.49 (H-2), 3.05 (H-3), 1.46 (H-5), 3.38 (H-6), 1.73,
1.45 (H-7), 1.77 (H-8), 1.76, 1.31 (H-11), 1.65, 1.60 (H-12), 2.11, 1.25 (H-15), 5.23 (H-16), 2.38 (H-17), 0.94 (H-18), 0.47,
0.17 (H-19), 1.24 (H-21), 2.13, 1.51 (H-22), 1.80, 1.69 (H-23), 3.61 (H-24), 3.81 (OH-25), 1.02 (H-26), 0.98 (H-27), 1.24
(H-28), 1.19 (H-29), 0.89 (H-30), 4.13 (H-1′-Xyl ), 2.96 (H-2′), 3.07 (H-3′), 3.24 (H-4′), 3.65, 3.00 (H-5′), 4.15 (H-1″-Glc ),
p
p
2.94 (H-2″), 3.11 (H-3″), 3.03 (H-4″), 3.05 (H-5″), 3.62, 3.44 (H-6″), 1.95 (CH COO).
3
13
C NMR spectrum (DMSO, δ, ppm, TMS): 31.32 (C-1), 29.08 (C-2), 87.16 (C-3), 41.47 (C-4), 51.19 (C-5), 77.27
(C-6), 32.57 (C-7), 44.03 (C-8), 20.38 (C-9), 27.95 (C-10), 25.51 (C-11), 31.88 (C-12), 45.31 (C-13), 46.66 (C-14), 44.33
(C-15), 74.95 (C-16), 56.69 (C-17), 19.44 (C-18), 27.33 (C-19), 84.78 (C-20), 26.86 (C-21), 36.32 (C-22), 25.69 (C-23), 82.08
(C-24), 70.11 (C-25), 26.79 (C-26), 24.87 (C-27), 19.72 (C-28), 27.22 (C-29), 15.78 (C-30), 105.98 (C-1′), 73.77 (C-2′), 76.59
(C-3′), 69.60 (C-4′), 65.50 (C-5′), 103.13 (C-1″), 74.02 (C-2″), 77.27 (C-3″), 70.22 (C-4″), 76.61 (C-5″), 61.29 (C-6″), 21.24,
171.25 (CH COO).
3
Alkaline Hydrolysis. Compound 1 (100 mg) was saponified by KOH solution (25 mL, 0.5%). The reaction mixture
was left at room temperature for 1 d, diluted with water (25 mL), and neutralized with acetic acid. The methanol was
evaporated. The solution was extracted with butanol. The solid after distilling off the butanol was chromatographed over a
silica-gel column with elution byCHCl :CH OH:H O (70:23:3) to afford 3 (70 mg) [6], C H O , mp 260-261°C (CH OH).
3
3
2
41 68 14
3
-1
IR spectrum (KBr, ν, cm ): 3382 (OH), 2941 (cyclopropane).
13
C NMR spectrum (C D N, δ, ppm, TMS): 34.66 (C-1), 29.03 (C-2), 88.27 (C-3), 42.67 (C-4), 52.56 (C-5), 79.20
5
5
(C-6), 34.92 (C-7), 46.24 (C-8), 21.13 (C-9), 28.93 (C-10), 26.49 (C-11), 33.41 (C-12), 45.08 (C-13), 46.24 (C-14), 45.77
(C-15), 73.41 (C-16), 58.23 (C-17), 21.13 (C-18), 30.23 (C-19), 87.27 (C-20), 28.60 (C-21), 32.24 (C-22), 26.20 (C-23), 81.69
(C-24), 71.29 (C-25), 28.24 (C-26), 28.60 (C-27), 27.10 (C-28), 16.65 (C-29), 19.87 (C-30), 107.56 (C-1′), 75.61 (C-2′), 78.15
(C-3′), 71.18 (C-4′), 67.07 (C-5′), 105.46 (C-1″), 75.61 (C-2″), 79.31 (C-3″), 71.29 (C-4″), 78.55 (C-5″), 63.19 (C-6″).
Acid Hydrolysis. Compound 3 (50 mg) was hydrolyzed for 3 h in methanolic H
SO (20 mL, 0.5%), cooled, and
2
4
treated with water (20 mL). The methanol was distilled off. The solution was heated for another 2 h, cooled, and extracted with
43

CHCl . The CHCl extracts were washed with water and evaporated to dryness in a rotary evaporator. The solid was
3
3
chromatographed over a silica-gel column with elution by CHCl :CH OH (9:1) to afford 4 (30 mg) [6, 7]. Paper
3
3
chromatographyusing butan-1-ol:pyridine:water (6:4:3) ofthe aqueous part ofthe hydrolysate detected D-xylose and D-glucose
by comparison with authentic specimens.
20
Literature data for 4 [6, 7]: mp 229-231°C (ethylacetate), [α]
+67.1° (c 0.192, CH OH).
3
D
Cyclostipuloside E (2). C H O , mp 272-274°C (CH OH).
53 94 24
3
-1
IR spectrum (KBr, ν, cm ): 3390 (OH), 2972 (cyclopropane).
PMR spectrum (500 MHz, C D N, δ, ppm, J/Hz, 0 = TMS): 0.25 and 0.45 (1H each, d, J = 3.9, 2H-19), 0.87, 0.97,
5
5
1.23, 1.35, 1.52, 1.68, 2.02 (s, 3H each, tertiary methyls), 3.64 (1H, dd, J = 11.6 and 4.6, H-3), 3.75 (1H, m, H-6), 4.78 (1H,
d, J = 7.6, xylose H-1′), 4.84, 4.92, and 5.20 (3H, d, J = 7.4, 7.6, and 7.8, glucose H-1″, H-1′″, and H-1″″).
13
For the C NMR spectrum, see Table 1.
Acid Hydrolysis. Glycoside 2 (40 mg) was hydrolyzed as described above to afford 5 (10 mg), C H O , mp 244-
30 52
5
23
246°C (acetone), [α]
+33.7 ± 2° (c 1.18, CH OH). These constants in addition to a comparison with an authentic specimen
3
D
identified the aglycon ascycloasgenin C(5)[2]. PCofthe hydrolysate using butan-1-ol:pyridine:water (6:4:3) detected D-xylose
and D-glucose by comparison with authentic specimens.
Enzymatic Hydrolysis. An aqueous solution of 2 (30 mg) was treated with H. plectotropis enzyme, held for 2 months
at 38 C, diluted with water (15 mL), and extracted with butanol (5 × 5 mL). The solvent was evaporated. The solid was
chromatographed over a silica-gel column with elution by CHCl :CH OH:H O (40:7.5:1) to afford progenin 6 (12 mg) [2],
3
3
2
C H O , mp 253-255°C (CH OH).
35 58
9
3
2
PMR spectrum (C D N, δ, ppm, J/Hz, 0 = TMS): 0.34 and 0.56 (1H each, d, J = 4.0, 2H-19), 0.99, 1.02 (d, J = 6.5),
5
5
3
1.25, 1.40, 1.47, 1.53, 1.94 (s, 3H each, tertiary methyls), 3.70 (1H, dd, J = 11.6 and 4.7, H-3), 3.74 (2H, m, H-6 and H-24),
3
4.86 (1H, d, J = 7.7, xylose H-1′).
13
For the C NMR, see Table 1.
Further elution of the column with the same solvent system isolated starting substance 2 (13 mg).
Acid Hydrolysis of Progenin 6. Compound 6 (12 mg) was hydrolyzed as described above to afford cycloasgenin C
(5, 4 mg). PC of the hydrolysate detected D-xylose by comparison with an authentic specimen.
ACKNOWLEDGMENT
We thank the leader of the NMR spectroscopygroup Dr. A. S. Shashkov and his coworkers for recording the PMR and
C NMR spectra.
13
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1.
T. N. Kaipnazarov, K. K. Uteniyazov, Z. Saatov, V. V. Kachala, and A. S. Shashkov, Khim. Prir. Soedin.,
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2.
3.
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448 (2001).
4.
T. N. Kaipnazarov, K. K. Uteniyazov, Z. Saatov, V. V. Kachala, and A. S. Shashkov, in: 5th International
Symposium on the Chemistry of Natural Compounds, Tashkent, 21-23 May, 2003.
E. Bedir, I. Calis, R. Aquino, S. Piacente, and C. Pizza, J. Nat. Prod., 62, 563 (1999).
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7.
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