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Boletín de la Sociedad Chilena de Química

versão impressa ISSN 0366-1644

Bol. Soc. Chil. Quím. v.47 n.4 Concepción dez. 2002

http://dx.doi.org/10.4067/S0366-16442002000400015 

SAPONINS FROM CARINIANA RUBRA (LECYTHIDACEAE)

EVANGELISTA LIMA1, PAULO TEIXEIRA DE SOUSA FILHO1,
JAIME BASTIDA2 AND GUILLERMO SCHMEDA-HIRSCHMANN3*

1Pós-Graduação em Saúde e Ambiente-ISC, Dep. Química I.C.E.T.,
Universidade Federal de Mato Grosso, Av. Fernando Correa da Costa s/n,
78060-900 Cuiabá, MT, Brasil.
2Universidad de Barcelona, Facultad de Farmacia, Departamento de Productos Naturales,
08028 Barcelona, Cataluña, España.
3*Laboratorio de Química de Productos Naturales, Instituto de Química de Recursos Naturales,
Universidad de Talca, Casilla 747, Talca, Chile.

(Received: April 15, 2002 - Accepted: July 24, 2002)

ABSTRACT

Three triterpenoidal saponins were isolated from the stem bark of Cariniana rubra Gardner & Miers (Lecythidaceae). The saponins reported are monodesmosides and a bidesmoside derivatives from the triterpene 2a ,3b ,23-trihydroxy-12-oleanen-28-oic acid (arjunolic acid). The proposed structures are 28-b -glucopyranosyl-23-O-acetyl arjunolic acid 1, 3-O-b -glucopyranosyl arjunolic acid 2 and 28-O-[ a -Rhamnopyranosyl-(1® 2)-b -glucopyranosyl] -23-O-acetyl arjunolic acid 3. The structures were elucidated mainly from its NMR spectral data. Compounds 1 and 3 are described for the first time.

Key Words: Cariniana rubra, Lecythidaceae, arjunolic acid derivatives, saponins.

RESUMEN

Tres saponinas triterpenoidales fueron aisladas de la corteza del tallo de Cariniana rubra Gardner & Miers (Lecythidaceae). Las saponinas que se informan son monodesmosidos y un derivado bidesmosidico del triterpeno ácido 2a ,3b ,23-trihidroxi-12-oleanen-28-oico (ácido arjunolico). Las estructuras propuestas son: ácido 28-b -glucopiranosil-23-O-acetil arjunolico 1, ácido 3-O-b -glucopiranosil arjunolico 2 y ácido 28-O-[ a -Rhamnopiranosil-(1® 2)-b -glucopiranosil] -23-O-acetil arjunolico 3. Los compuestos 1 y 3 se describen por primera vez.

PALABRAS CLAVES: Cariniana rubra, Lecythidaceae, derivados del ácido arjunolico, saponinas.

INTRODUCTION

Several trees belonging to the plant family Lecythidaceae are known under the common name "jequitibá" in Brazil. The name "jequitibá" is usually accompanied by a distinctive characteristic of the tree, usually the wood or flower color. The large tree Cariniana rubra Gardner & Miers is known as "jequitibá vermelho"1). A bark infusion is used in the Mato Grosso traditional medicine as antiinflammatory.

According to Hegnauer2), triterpene, triterpensaponine as well as tannins are widespread in the Lecythidaceae. However, little is known on the chemistry of the genus Cariniana. The aim of this work was to isolate and determine the structure of the saponins from C. rubra stem bark.

EXPERIMENTAL

a. Plant material

The plant material was collected at the Vale do Sol, nearly Rio Formosa, Cuiabá, MT. A voucher herbarium specimen (Exsicata Nº 10.096) has been deposited at the Herbario Central da UFMT. The plant material was dried at 25ºC, ground and extracted successively with hexane and methanol to yield a crude hexanic and a methanolic extract. Preliminary assays3) indicated the occurrence of tannins, saponins, steroids, triterpenes and flavonoids in the extracts.

b. Isolation

The methanolic extract of "Jequibibá vermelho" stem bark was extracted with EtOAc under reflux to afford an EtOAc-soluble (5.5%) and an EtOAc-insoluble fraction. The EtOAc-soluble portion (5.5 g) was chromatographed in a medium pressure column (Silica gel) with a petroleum ether (PE) ¾ PE:EtOAc ¾ EtOAc gradient to obtain 20 mg stigmasterol, 15 mg b -sitosterol, 30 mg b -amyrine and 50 mg of the 3-O-b -D-glucoside of sitosterol4-5). Some 10 g of the EtOAc-insoluble portion from the MeOH extract was permeated on Sephadex LH 20 with MeOH. Fractions were collected as follows: 200 fractions of 30 mL each, 24 fractions of 220 mL and an acetone fraction of 800 mL. After TLC comparison fractions with similar TLC patterns were pooled resulting in nine groups. The supernatant of fraction 2 from the Sephadex column (446 mg) showed positive reaction to saponins and was purified by high speed circular countercurrent chromatography (CCCC) using the following system: CHCl3: MeOH: H2O (13:7:8). The lower phase was used as the mobile phase6). Some 100 fractions of 5 mL each were colected. After TLC comparison fractions were pooled to afford 8 mg 1, 6 mg 2 and 11 mg 3.

00000Compound 1 (28-b -glucopyranosyl-23-O-acetyl arjunolic acid): white powder, EI MS (m/z, rel. int. %): 248 (100) (C16H24O2); 203 (78) (248-CO2H). IR (KBr) cm-1: 3340, 1750, 1640, 885, 1000-1100.

00000Compound 2 (3-O-b -glucopyranosyl arjunolic acid): colorless powder, EI MS (m/z, rel. int. %): 248 (100) (C16H24O2); 203 (80) (248-CO2H). IR (KBr) cm-1: 3400, 1740, 1000-1100.

00000Compound 3 (28-O-[ a -Rhamnopyranosyl-(1® 2)-b -glucopyranosyl] -23-O-acetyl arjunolic acid) : colorless powder, EI MS (m/z, rel. int. %): 248 (100) (C16H24O2); 203 (85) (248-CO2H). IR (KBr) cm-1: 3400, 1750, 1000-1100.

Results and discussion

Three saponins were isolated from the EtOAc-insoluble fraction of a methanolic extract of Cariniana rubra stem bark. The EI-MS of the three compounds showed strong signals at m/z 248 and 203, characteristic of a retro Diels-Alder fragmentation of olean-12-en-28-oic or ursan-12-en-28-oic acid derivatives without additional substitution in rings C, D and E 7).

The 1H-NMR spectrum of the genine in compounds 1-3 showed six methyl singlets and a CH2OH group, a dd at d 2.85-3.18 and a broad singlet at d 5.38-5.48 suggesting a12-oleanen-28-oic acid derivative. The EI-MS supported this assumption. The presence of two hydroxy functions in the A ring followed from the d at d 3.03-3.81 (j = 9.3¾9.5 Hz) coupling with a second signal at d 3.77-4.30 ppm. Two-dimensional H-H- spectra clearly shows a -CHOH-CHOH-CH2- sequence with coupling constants in agreement for a 2a , 3b configuration8). The 13C NMR spectra of the genine in the compounds 1, 2 and 3 pointed out to 2,3,23-trihydroxy-12-oleanen-28-oic acid derivatives with glycosidation at C-3 or C-28 with or without acetylation at C-23. The 13C NMR data for the genine is in agreement with those reported for 2a , 3b , 23-trihydroxi-28-oic acid (arjunolic acid)4,6,9-10).

The placement of the glycosidic moiety can be deduced from the chemical shift of the C-3 and C-28 signal in the 13C NMR spectra. Typical shifts for unsubstituted C-3 and C-28 positions ranges between 73-79 and 180.1-180.4 ppm while upon glycosidation the values are 82.8-86.7 and 176.3-176.6 ppm, respectively10-11). In ursane derivatives, Abe and Yamauchi12) report values of 68.7 and 81-84 ppm for C-2 and C-3 in unsubstituted dihydroxy compounds, respectively.

The molecular formula of compound 1 follows from the 13C data which indicates a molecular formula of C38H60O11 and pointed out to a monodesmosidic derivative of 23-acetyl arjunolic acid. The 1H-NMR spectrum showed a anomeric proton at d 6.28 (j = 7.9 Hz) consistent with a b -configurated sugar unit. The 13C-NMR spectra allowed the identification of the sugar as glucose. Hydrolisis of compound 1 and co-chromatography with standards afforded glucose as the sugar component. The chemical shift of the C-28 signal in the 13C-NMR spectrum (d 176.62) indicates that glycosidation took place at C-28. The same position for arjunolic acid resonates at d 180.0 ppm. No significant changes were observed for the chemical shifts of C-2 and C-3 signals in arjunolic acid compared with compound 1. The structure was assigned to 28-b -glucopyranosyl-23-O-acetyl arjunolic acid.

The 1H-NMR spectrum of the compound 2 was close to that of 1 differing in the absence of the acetate signal and showing a d at d 6.31 (j = 7.9 Hz) indicating a b -configurated sugar. In the 13C-NMR spectrum, the chemical shift of C-3 from 77.36 to 85.55 ppm clearly indicates that glycosidation took place at C-3. The sugar was identified as glucose by its NMR spectra. and allowed the identification of the compound as 3-O-b -glucopyranosyl arjunolic acid.

The NMR data of the compound 3 pointed out to an oleanene saponin with two sugar units. The anomeric protons indicates the presence of two monosaccharide units, one of them belonging to rhamnose and a b -configurated glucose. Both 1H and 13C-NMR data for the sugar moiety are in complete agreement with the data reported for a -rhamnopyranosyl-(1® 2)-b -glucopyranosyl in Asteryuannanoside A9) Based on the spectroscopic data we propose for 3 the structure 28-O-[ a -Rhamnopyranosyl-(1® 2)-b -glucopyranosyl] -23-O-acetyl arjunolic acid. The 1H and 13C-NMR data of the compounds 1, 2 and 3 are presented in Tables I ¾ III.

Table I. 1H-NMR data of compounds 1-3 (300 MHz, pyridine-d5 for 1-3, 500 MHz, MeOH-d4 for 2-3, d H', ppm).


H

1 (Pyr)

2 (Pyr)

2 (MeOH)

3 (Pyr)

3 (MeOH)


1

 

1.1 m
2.2 m

0.85 m
1.94 m

 

0.8 m
1.9 m

2

4.25 m

4.30 m

3.77 ddd
(11; 9.5; 4)

4.20 m

3.66 m

3

3.81 d (9.3)

3.55 d (9.5)

3.03 d (9.5)

 

3.26 d (9.5)

11

 

1.9 m

1.92 m

2.04 m

1.95 m (2H)

12

5.38 br s

5.40 br s

5.25 dd (3.5; 4)

5.48 br s

5.28 dd (3.5; 3)

16

   

2.04 ddd
(13.5; 13.5; 4)

   

18

3.18 dd
(13.5; 4)

3.15 dd
(13.5; 4)

2.85 dd
(13.5; 4)

3.12 dd
(13.5; 4)

2.82 dd
(13.5; 4)

19

 

1.2 m
1.6 m

1.16 m
1.70 m

1.20 m
1.8 m

1.18 dd (14, 4)
1.72 dd (14, 13.5)

       

1.33 t

 

Methyl

0.84 s

0.86 s

0.77 s

0.76 s

0.76 s

 

0.84 s

0.88 s

0.90 s

0.82 s

0.81 s

 

0.97 s

1.00 s

0.92 s

0.95 s

0.90 s

 

1.00 s

1.09 s

0.98 s

1.04 s

0.93 s

 

1.09 s

1.20 s

1.15 s

1.11 s

1.04 s

 

1.19 s

1.57 s

1.22 s

1.21 s

1.15 s

OAc

1.88 s

-

-

1.89 s

2.04 s

23

   

4.01 d (11)
3.33 d (11)

3.84 d (9.3)
4.12 d (9.3)

3.95 d (11)
3.92 d (11)

Glucose

         

1

6.28 d (7.9)

6.31 d (7.9)

5.37 d (8)

6.22 d (8)

5.43 d (8)

2

3.95-4.45 m

4.18 t (7.9)

3.31 m

4.51 dd
(7, 4.3)

3.54 dd (9; 8)

3

3.95-4.45 m

4.4 m

 

4.38 m

3.38 dd (9.5)

4

3.95-4.45 m

3.68 m

 

4.35 m

 

5

3.95-4.45 m

4.0 m

3.36 m

3.98 m

3.33 m

6

3.95-4.45 m

4.2 m

3.67 dd (12, 4.5)
3.79 dd (11, 2)

4.3 m

3.78 dd
3.66 dd

Rhamnose

         

1

-

-

-

6.63 br s

5.37 d (1.5)

2

-

-

-

4.82 d (1.1)

3.58 m

3

-

-

-

4.56 m

 

4

-

-

-

4.35 m

 

5

-

-

-

4.56 m

3.74 dq (9; 6.5)

6

-

-

-

1.76 d (6.5)

1.23 d (6.5)


Table II. 13C NMR data of the aglycone moieties of compounds 1-3 (75 MHz, pyridine-d5 for arjunolic acid and 1, MeOH-d4 for 2 and 3, d C, ppm).


C

1

2

3

Arjunolic acid


1

47.16 t

48.00

47.50

47.6

2

68.56 d

69.56

70.25

68.8

3

77.36 d

85.88

79.23

78.2

4

42.98 s

44.41

43.49

43.6

5

48.57 s

57.15

54.20

48.0

6

18.80 t

19.86

19.26

18.5

7

30.14 t

33.49

33.52

32.9

8

40.18 s

40.70

40.78

39.8

9

48.57 s

48.00

48.16

48.1

10

38.45 s

39.10

39.02

38.4

11

23.55 t

24.87

24.66

23.7

12

122.92 d

123.48

123.41

123.2

13

144.34 s

144.92

144.91

144.9

14

42.28 s

42.92

42.89

42.2

15

29.00 t

28.85

29.20

28.3

16

24.09 t

23.78

24.15

23.9

17

46.29 s

47.21

47.20

47.6

18

41.92 d

42.57

42.90

42.0

19

39.51 t

40.71

43.05

46.4

20

30.91 s

30.77

31.57

30.9

21

34.14 t

33.13

34.91

34.1

22

32.68 t

31.56

32.98

33.2

23

67.00 t

66.19

66.96

66.6

24

14.08 q

17.7

13.85

14.2

25

17.56 q

17.63

17.70

17.3

26

17.74 q

17.63

17.77

17.5

27

26.08 q

26.29

26.12

26.1

28

176.62 s

178.01

177.90

180.0

29

33.24 q

34.16

33.50

33.2

30

23.80 q

23.97

24.10

23.7

OAc

20.84 q

-

20.83

-

 

170.87 s

-

172.69

-


Table III. 13C NMR data of the glycoside moieties of compounds 1-3 (75 MHz; pyridine-d5 for 1, MeOH-d4 for 2 and 3, d C, ppm).


 

1

2

3


Glc 1

95.95

95.68

95.08

Glc 2

74.29

73.89

77.07

Glc 3

79.09

78.29

78.43

Glc 4

71.29

71.08

71.30

Glc 5

79.50

78.70

77.71

Glc 6

62.04

62.38

62.36

Rh 1

-

-

101.57

Rh 2

-

-

71.93

Rh 3

-

-

72.18

Rh 4

-

-

73.69

Rh 5

-

-

69.31

Rh 6

-

-

18.24


Conclusions

Arjunolic acid has been reported as a constituent of Terminalia arjuna, Tristania conferta, Prunella vulgaris, Polygala japonica, Metrosideros umbellata, Psidium guajava and Mitragyna ciliata. Its b -D-glucopyranosyl ester has been isolated from Tristania arjuna and was described as Arjunglucoside II. Closely related compounds were also obtained from T. arjuna. Arjunolitin is the (3-O-b -D-glucuronopyranoside, b -D-glucopyranosyl ester) from Tristania arjuna. The [ a -L-Rhamnopyranosyl-(1® 2)-b -D-glucopyranosyl] ester of arjunolic acid, known as Asteryuannanoside A was first reported from Aster yunnanensis 9,13).

The new saponins reported are monodesmosides and bidesmoside derivatives from the triterpene 2a ,3b ,23-trihydroxy-12-oleanen-28-oic acid (arjunolic acid) and its 23-acetate. The proposed structures are 28-b -glucopyranosyl-23-O-acetyl arjunolic acid 1, 3-O-b -glucopyranosyl arjunolic acid 2 and 28-O-[ a -Rhamnopyranosyl-(1® 2)-b -glucopyranosyl] -23-O-acetyl arjunolic acid 3. The isolation of arjunolic acid 3-O-b -D-glucopyranoside from Dolichos kilimandscharicus has been reported10). Related compounds have been reported from Terminalia arjuna14). Compounds 1 and 3 are reported for the first time.

ACKNOWLDGMENTS

We are grateful to IFS (Grant F 2491/1 to Paulo T. de Sousa Jr.), CNPq and SUDAM (Brazil) and the Programa de Productos Bioactivos, Universidad de Talca (Chile) for financial support.

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*To whom correspondence should be addressed.