Triterpene and steroidal glycosides of the genus Melilotus and their genins. VI. Melilotoside A2 and adzukisaponin v from the roots of M. tauricus

Article abstract of DOI:10.1007/s10600-013-0455-x

Two triterpene glycosides of the oleanane series, melilotoside A ₂ and previously known adzukisaponin V, were isolated from the roots of Melilotus tauricus (L.) Pall. (Fabaceae). The structures of the glycosides were proved based on chemical transformations. Melilotoside A₂ had the structure soyasapogenol B 3-O-β-D-glucouronopyranoside; adzukisaponin V, soyasapogenol B 3-O-[α-L-rhamnopyranosyl-(1-2)-β-Dglucopyranosyl- (1-2)-β-D-glucuronopyranoside].

Full text of DOI:10.1007/s10600-013-0455-x

Chemistry of Natural Compounds, Vol. 48, No. 6, January, 2013 [Russian original No. 6, November–December, 2012]
TRITERPENE AND STEROIDAL GLYCOSIDES
OF THE GENUS Melilotus AND THEIR GENINS.
VI. MELILOTOSIDE A AND ADZUKISAPONIN V
2
FROM THE ROOTS OF M. tauricus
G. V. Khodakov
UDC 547.918:582.738:543.422
Two triterpene glycosides of the oleanane series, melilotoside A and previously known adzukisaponin V,
2
were isolated from the roots of Melilotus tauricus (L.) Pall. (Fabaceae). The structures of the glycosides
were proved based on chemical transformations. Melilotoside A had the structure soyasapogenol B 3-O-ꢀ-
2
D-glucouronopyranoside; adzukisaponin V, soyasapogenol B 3-O-[ꢁ-L-rhamnopyranosyl-(1ꢂ2)-ꢀ-D-
glucopyranosyl-(1ꢂ2)-ꢀ-D-glucuronopyranoside].
Keywords: Melilotus tauricus, Fabaceae, adzukisaponin V, triterpene glycoside melilotoside A .
2
We have proved the structures of triterpene glycosides from the roots of Melilotus tauricus growing in Crimea.
Chemical structures of a new triterpene glycoside called melilotoside A (1) and the previously known adzukisaponin V (2)
2
that was identified by us were established.
An analysis of total acid hydrolysis products of each studied compound using PC and TLC (characterized by comparison
of their chromatographic mobilities with authentic standards) showed the presence in each of the aglycon soyasapogenol B (3)
(3ꢀ,22ꢀ,24-trihydroxyolean-12-ene). The aglycon was present in the native form in the CHCl extract of the root extract of
3
this plant.
13
The carbohydrate part of melilotoside A was glucuronic acid. This was confirmed by the presence in the C NMR
2
ꢂ1
spectrum of the corresponding resonances (Table 1) [1]; in the IR spectrum, a band at 1740 cm corresponding to a carboxylic
C=O [2]. A comparison of the chemical shifts of glucoside C atoms in the unsubstituted portions with the literature showed
that the carbohydrate constituent was bonded to C-3 of the aglycon (weak-field shift from 80.2 to 91.3 ppm [3]).
Partial acid hydrolysis of 2 and chromatographic separation of the hydrolysis products isolated two progenins (2a and
2b). The TLC chromatographic mobility of progenin 2a coincided with that of 1. Total acid hydrolysis of the progenins and
the glycoside showed according to PC the presence of glucuronic acid in 2a and glucuronic acid and glucose in 2b whereas
glycoside 2 contained glucuronic acid, glucose, and rhamnose. Thus, the chemical structure of the glycoside sugar component
consisted of sequential alternation of glucuronic acid, glucose, and rhamnose. Furthermore, progenin 2a was identical to 1
according to chemical transformations, TLC, and PC.
The bond types in 2 were established by GC of the methanolysis products of the glycoside permethylate that was
prepared by Hakomori methylation [4] and subsequent identification of methyl-3,4,6-tri-O-methyl-ꢀ-D-glucuronopyranoside;
methyl-3,4,6-tri-O-methyl-ꢀ-D-glucopyranoside; and methyl-2,3,4-tri-O-methyl-ꢁ-L-rhamnopyranoside in the ratio
0.98:1.04:1.02.
Thus, we propose for melilotoside A the structure soyasapogenol B 3-O-ꢀ-D-glucuronopyranoside. Glycoside 2
2
was identified as adzukisaponin V {soyasapogenol B 3-O-[ꢁ-L-rhamnopyranosyl-(1ꢂ2)-ꢀ-D-glucopyranosyl-(1ꢂ2)-ꢀ-D-
glucuronopyranoside]}.
Southern Filial, National University of Bioresources and Nature Utilization of Ukraine, Crimea Agrotechnology
University, s. Agrarnoe, 95492, Simferopol, Crimea AR, Ukraine, e-mail: commission@csau.crimea-ua.com. Translated from
Khimiya Prirodnykh Soedinenii, No. 6, November–December, 2012, pp. 906–907. Original article submitted April 7, 2012.
©
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0009-3130/13/4806-1024 2013 Springer Science+Business Media New York

13
TABLE 1. C NMR Chemical Shifts for Melilotoside A (1) (C D N, 0 = TMS, ꢄ, ppm)
2
5 5
C atom
C atom
C atom
ꢄÑ
ꢄÑ
ꢄÑ
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
38.5
26.6
91.3
44.5
56.0
19.3
33.7
40.1
47.5
36.8
24.24
122.3
144.5
42.7
26.9
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
38.5
38.6
45.3
46.7
31.4
42.6
75.3
23.3
63.6
15.8
17.4
25.7
26.5
33.5
21.9
ꢀ-D-Clur
p
107.6
80.1
76.6
75.4
76.1
171.9
1
2
3
4
5
6
OH
RO
1 - 3
OH
1: R = ꢀ-D-Glurꢅ
2
2
2: R = ꢁ-L-Rhaꢅ ꢀ-D-Glcꢅ ꢀ-D-Glurꢅ
3: R = H
EXPERIMENTAL
General Comments. The used solvent systems; TLC, CC, and PC conditions; and spectral analyses were described
before [5]; GC, in [6].
Isolation of Glycosides. Ground air-dried roots of M. tauricus (1 kg) that were collected during flowering were
extracted with aqueous EtOH (70%). The extract was evaporated to one third the volume and extracted successively with
CHCl and BuOH. The BuOH extract was evaporated to afford total extracted substances (15 g). Chromatography used
3
systems 1, 2, and 3 sequentially to afford fractions enriched in 1 and 2.
Melilotoside A and Adzukisaponin V (1, 2). Fractions enriched in glycosides 1 and 2 were evaporated to dryness.
2
The residues were dissolved in aqueous KOH (5%), refluxed on a water bath for 2 h, diluted with H O (to 100 mL), and
2
extracted with BuOH. The extracts were evaporated to dryness and chromatographed over SiO with elution by systems 1 and
2
3 for 1 and 2, respectively, to afford 0.015 g (0.002% of air-dried raw material mass) of 1, C H O , mp 220–221°C (dec.)
36 57
8
ꢂ1
(from system 1), IR spectrum (KBr, ꢃ, cm ): 3580-3200 (OH), 1740 (carboxylic C=O) and 0.05 g (0.006%) of 2, C H O ,
mp 225–227°C (dec.) (from system 2), IR spectrum (KBr, ꢃ, cm ): 3580–3200 (OH), 1720 (carboxylic C=O).
48 78 18
ꢂ1
Total Acid Hydrolysis of the Glycosides. Glycosides 1 and 2 were dissolved in aqueous H SO (10 mL, 5%),
2
4
refluxed on a sand bath for 24 h, diluted with H O, and extracted several times with CHCl . The combined extract was washed
2
3
with H O until neutral and evaporated to dryness. The residue was chromatographed over silica gel using system 4 to afford
2
the genins of 1 (4 mg) and 2 (5 mg). These were identified as soyasapogenol B according to mp and TLC.
The hydrolyzate was neutralized with BaCO , evaporated to dryness, and dissolved in the minimum volume of
3
MeOH. PC using system 6 identified glucuronic acid in 1 and glucose, glucuronic acid, and rhamnose in 2.
Partial Acid Hydrolysis of 2. Glycoside 2 (20 mg) was dissolved in aqueous H SO (15 mL, 2.5%), refluxed on a
2
4
sand bath for 4 h, diluted with H O, and extracted several times with water-saturated BuOH. The combined BuOH extract was
2
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washed until neutral and evaporated to dryness. Chromatography using successively systems 4, 1, and 2 isolated progenin 2a
(5 mg), mp 220–221°C (dec.) (system 1) and progenin 2b (7 mg), mp 222–224°C (dec.) (system 2).
Total Acid Hydrolysis of Progenins 2a and 2b. The progenins (4 and 6 mg, respectively) were dissolved in aqueous
H SO (5 mL, 5%) and refluxed on a sand bath for 24 h. The mixtures were neutralized with BaCO , filtered, evaporated to
2
4
3
dryness, and dissolved in the minimum volume of MeOH. PC using system 6 identified glucuronic acid in 2a and glucuronic
acid and glucose in 2b.
Permethylate of 2. Glycoside 2 (50 mg) in anhydrous DMSO (10 mL) was stirred, treated in small portions with
NaH (50 mg) and dropwise with MeI (2 mL), stirred for another 5 h, and poured into aqueous sodium hyposulfite (50 mL, 2%).
The products were extracted with CHCl . The usual work up of the CHCl extract and evaporation of solvent afforded a
3
3
residue that was chromatographed over a column with elution by system 5 to isolate the methanolysis products.
GC of the methanolysis products detected methyl-3,4,6-tri-O-methyl-ꢀ-D-glucuronopyranoside, methyl-3,4,6-tri-O-
methyl-ꢀ-D-glucopyranoside, and methyl-2,3,4-tri-O-methyl-ꢁ-L-rhamnopyranoside in the ratio 0.98:1.04:1.02.
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