ISOLATION AND STRUCTURE ELUCIDATION OF A NEW OLEANANE TYPE GLYCOSIDE FROM THE AERIAL PORTION OF CESTRUM NOCTURNUM

ABSTRACT. Cestrum nocturnum (Solanaceae) is an ornamental plant cultivated in various parts of the world due to its sweet-scented white flowers. It is commonly called night-blooming Jessamine (Raat ki Rani). The genus is known for its toxicity to feedents. The leaves may cause uneasiness in animals which may lead to severe gastroenteritis. The plant is known to be a rich source of pharmacologically active saponins. Looking to its various pharmacological activities as reported, the plant was explored for the isolation of new phytochemicals. During the process, a new oleanen type glycoside was isolated from the butanolic fraction of the leaves of Cestrum nocturnum and was characterized as 3-O-β-D-xylopyranoside-olean-12-en-28-oic acid-28-O-β-arabinopyranosyl(1-3)-β-D-galacto-pyranosyl-(1-2)-β-D-glucopyranosyl-(1-4)-β-D-glucopyranosyl ester, along with two reported compounds nocturnoside A and karativoside A. The structure was elucidated on the basis of 1D and 2D NMR and mass spectrometry.


INTRODUCTION
Cestrum is an important genus of the Family Solanaceae, native to tropical America and Australia and is cultivated in various parts of the world for ornamental purpose due to their sweet-scented white flowers [1]. The genus is known for its toxicity to feedents. The species of this genus have cymose inflorescences that are terminal in position or born axillary and are usually fragrant. There are 175 species belonging to genus Cestrum that are native to tropical America and Australia. In Pakistan four cultivated species of the genus Cestrum exists, that includes C. aurantiacum, C. diurnum, C. nocturnum, and C. parqui [2]. They are commonly known as Cestrum or jessamine due to their fragrant flowers. Cestrum nocturnum is commonly called night-blooming jessamine (Raat ki Rani) [2][3][4]. Cestrum nocturnum leaf and flower, if ingested cause uneasiness in animals which may lead to severe gastroenteritis and is larvicidal [5]. The toxicity was assigned to the derivatives of tigogenin, tigonin [6,7] and ursolic acid [8]. The essential oil has demonstrated high disease inhibition efficiency on greenhouse-grown pepper plants [9]. The antimicrobial potential was also reported [10]. Alkaloids, saponins, phenolic compounds, tannins, and flavonoids are reported from the species [6]. Some interesting and high molecular weight saponins have been isolated from the species. Phytochemicals from this genus exhibit wide range of pharmacological significances in skin disorders and treating arterial hypotension. They are also used as antiviral, analgesic, abortive, diuretic, antispasmodic, dyspeptic, smooth muscle relaxant, negative inotropic and chronotropic agent [11][12][13].

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In the course of phytochemical investigations, we isolated a new triterpene glycoside, along with two reported glycosides nocturnoside A [14] and karativoside A [15]. The new structure is a bisdesmosidic glycoside of oleanolic acid [16] with one sugar unit attached at C-3 and other oligosaccharide units to C-28.

RESULTS AND DISCUSSION
A white amorphous powder (32 mg) was isolated from iso-butanol fraction of Cestrum nocturnum and its molecular formula was determined as C 58  The oleanane-type triterpenoid saponin nature of the isolated compound was revealed through analysis of its spectral features. Bisdesmosidic nature of the glycoside was established due to the upfield chemical shift at C-28 and the absence of the distinguished carboxyl proton signal at δ H 10-12, in the 1 H-NMR spectrum [19].
The assignments for the vinylic H-12 proton at δ H 5.32 was confirmed by its COSY-45 o correlations with H-11α (δ H 1.78) and H-11β (δ H 1.83), and in turn the protons at C-11 are showing coupling with H-9 (δ H 1.28). The assignments of protons at position 19 were confirmed by their interactions with H-18α (δ H 4.40) and H 3 -29 (δ H 0.94) [19].
In For the identification of monosaccharide, acid hydrolysis of the compound was performed with 20% HCl. The sugars were identified as xylose, arabinose, galactose and glucoe on the basis of paper chromatography using ninhydrin as visualizing agent [1].
Two other compounds nocturnoside A and karativoside A were also isolated from the plant. All the spectroscopic techniques were applied and the structures of were confirmed by comparing the spectra with the reported data ( Figure 2).

EXPERIMENTAL
Cestrum nocturnum leaves were shade dried crushed and then extracted with methanol. The dried methanolic extract was partitioned with various solvents having different polarities, including, n-hexane, ethyl acetate and iso-butanol. The butanolic fraction was passed through vacuum liquid chromatography, using ethyl acetate-hexane (EtAc/Hex) and then ethanol and ethyl acetate (EtOH/EtAc) mixed in various proportions and different fractions were collected from 17-45.