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ISOLATION AND CHARACTERIZATION OF THE CHEMICAL CONSTITUENTS OF STELLARIA MEDIA LINN.

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ISOLATION AND CHARACTERIZATION OF THE CHEMICAL CONSTITUENTS OF STELLARIA MEDIA LINN.

Disha Arora and Anupam Sharma *

University Institute of Pharmaceutical Sciences, Panjab University, Chandigarh - 160014, Punjab, India.

ABSTRACT: The studies on the constituents of aerial parts of Stellaria media Linn. (Caryophyllaceae) have led to the isolation and characterization of three new compounds ─ (1), (2), and (3). Structure elucidation of the new compounds was carried out using 1H NMR, 13C NMR, Mass spectrometry, together with other complementary techniques (UV and IR). The major findings of the study include: Compound 1 was separated as a white amorphous solid from petroleum ether extract of S. media, Methanol extract of S. media was subjected to bioactivity-guided fractionation, and Compounds 2 and 3 were isolated, and characterized by different spectral studies. The interpretation of the three compounds include: compound 1 interpreted as 2, 4, 5, 7-tetramethyloctane, compound 2 as - 6-methyl heptyl-3′-hydroxy-2′-methyl propanoate; and compound 3 as - 2, 2, 4-trimethyloctan-3-one.

Keywords:

Stellaria media, Caryophyllaceae, 2, 4, 5, 7-tetramethyloctane, 6-methyl heptyl-3′- hydroxy-2′-methylpropanoate, 2, 2, 4-trimethyloctan-3-one

INTRODUCTION: Plants of the genus Stellaria (Caryophyllaceae) are annuals or perennials, distributed in the cold and temperate regions. The genus consists of about 120 species of flowering plants of the family Caryophyllaceae 1. About 23 species occurring in India 2 are reputed for their medicinal properties. Among these, S. media seems to possess wide medicinal potential. The plant has been used traditionally in the treatment of mental tension and inflammations of the digestive, renal, respiratory, and reproductive tracts 3. The plant is also reported to be anticancer 4, antipyretic, anti-inflammatory 5, antibacterial, antifungal 6, 7, anxiolytic 8.

Phenolic acids, flavonoids 9, 10, triterpenoid saponins 11, a pentasaccharide 12, lipids 13 and aqueous constituents 14 have been reported from the genus Stellaria. The study described in this paper was, therefore, undertaken to further examine the phytochemical profile of this plant. Bioactivity-guided fractionation of the anxiolytic methanol extract has led to the isolation of three compounds 1-3. Structures of the compounds were established by spectral studies.

MATERIALS AND METHODS:

General Experimental Procedures: TLC plates (silica gel 60 F254) were purchased from Merck (Darmstadt, Germany). Column chromatography was carried out on silica gel 60, E. Merck, mesh size 60-120, and 230-400. The following instruments at National Institute of Pharmaceutical Education and Research, Mohali were used: IR spectrophotometer (Multi spoke FT-IR synthesis monitoring system, Perkin-Elmer, Germany); mass spectrometer (Finnigan, MAT, LCQ, USA) equipped with a pneumatically-assisted atmospheric-pressure chemical ionization (APCI). 1H NMR and 13C NMR spectra were obtained on a 400 MHz NMR spectrometer, (Bruker 400, Ultra Shield, ZH079807, Avane, Germany) using CDCl3 as a solvent. Chemical shifts were expressed in parts per million (ppm) relative to tetramethyl silane (TMS) as an internal standard. All solvents used for plant extraction and crude separations were of analytical grade.

Plant Material: Aerial parts of S. media were collected from around the University Institute of Pharmaceutical Sciences building, Panjab University, Chandigarh in February 2009. Identity of the plant was confirmed by Dr. H.B. Singh, Head, Raw Materials, Herbarium & Museum at the National Institute of Science Communication and Information Resources, (NISCAIR, CSIR), New Delhi 110 067. A voucher specimen no: NISCAIR/RHMD/Consult/2008-09/1170/202 is deposited in the same herbarium.

Extraction and Isolation: Scheme 1 shows the extraction of S. media aerial parts, and isolation of three compounds (1-3).

SCHEME 1: ISOLATION OF COMPOUNDS 1-3

Powdered aerial parts of the plant (1 kg) were subjected to successive Soxhlet extraction by solvents in increasing order of polarity viz. petroleum ether (60-80 °C), chloroform and methanol. From the concentrated petroleum ether extract (2 g) of S. media, an amorphous material separated after about 48 h, and this was collected by filtration. The residue on the filter paper was washed (3 × 2 ml) with petroleum ether, and finally, air dried to get 30 mg of an amorphous solid.

TLC of the separated material using cyclohexane: toluene (9.5:0.5) showed a single spot (Rf 0.95) when visualized using 0.5% anisaldehyde sulphuric acid spraying reagent. This compound was labeled as 1. Earlier studies carried out by the authors revealed that of the four extracts, only the methanol extract exhibited anxiolytic activity 8. Repeated bioactivity guided fractionation of methanol extract using solvent partitioning and column chromatography led to the separation of a subfraction EAF5 (Ethyl acetate fraction 5) which was found to be responsible for the antianxiety activity of the methanol extract 15. The present study was carried out to separate the components in EAF5. The TLC profile of EAF5 showed the presence of two components (Rf 0.73 and 0.78, respectively). These two compounds were separated using preparative TLC. From 150 mg of EAF5, two compounds were obtained, namely 2 (75 mg) and 3 (50 mg).

Characterization of 2, 4, 5, 7-tetramethyloctane (1): White amorphous solid; m. p. 48-50 °C; UVmax (MeOH) 180 nm; IR (KBr) 722.9, 1376.2, 1461.9, 2852.5, 2920.8 cm-1;   1H-NMR (CDCl3) δ 0.84 (br, 4H, 2 x -CH2), 0.86 (br, 2H, 2 x -CH), 1.25 (br, 18H, 6 x -CH3), 1.55 (m, 2H, 2 x -CH), ppm; 13C-NMR (CDCl3) δ 14.09 (2 × -CH-CH3), 22.67 [2 × -CH (CH3)2], 29.68 [2 × -CH (CH3)2], 31.92 (2 ×-CH-CH3), 37.10 (-CH2) ppm; MS (APCI) m/z 170 [M]+ (C12H26), 172 [M+2]+, 127 [M - CH (CH3)2]+, 85 [M – CH (CH3) (CH2)-CH (CH3)2]+, 57 [M – CH (CH3)-CH (CH3)-CH2-CH (CH3)2]+.

Characterization of 6-methylheptyl-3′-hydroxy-2′-methylpropanoate (2): Yellow semi-solid; m. p. 70-72 °C; UVmax (MeOH) 246 nm; IR (KBr) 1090.7, 1157.4, 1262.1, 1378.1, 1458.4, 1727.6, 2857.0, 2924.2, 3426.0 cm-1; 1H-NMR (CDCl3) δ 0.80 (m, 2H, -CH2), 1.08 [br, 1H, -CH (CH3)2], 1.35 (br, 6H, 3 × -CH3), 1.52 (m, 2H, -CH2), 1.72 [br, IH, -CH (CH2OH) (CH3], 1.83 (m, 2H, -CH2), 2.07 (m, 2H, -CH2), 2.35 (d, 3H, -CH3), 2.70 (br, 1H, OH), 4.32 (t, 2H, -CH2) ppm; 13C-NMR (CDCl3) δ 14.13 [-CH (CH3) (CH2OH)], 22.70 [-CH (CH3)2], 25.52 [-CH2-(CH2)2-CH (CH3)2], 27.09 [-CH2-CH2-CH (CH3)2], 29.36 (-CH (CH3)2), 29.66 (-CH2-CH2-O-C=O), 37.09 (-CH2-CH (CH3)2), 50.04 (-CH (CH3) (CH2OH), 57.36 (-CH2OH), 61.70 (-CH2-O-), 185.56 (-C=O) ppm; MS (APCI) m/z 216 [M]+ (C12H24O3), 217 [M+1]+, 173 [M – CH (CH3)2]+, 145 [M – CH (CH3)2-(CH2)2]+, 131 [M  – CH (CH3)2-(CH2)3]+, 103 [M – O-C=O-CH (CH3)-CH2OH]+.

Characterization of 2, 2, 4-trimethyloctan-3-one (3): White crystals; m. p. 76-78 °C; UVmax (MeOH)  243 nm; IR (KBr) 1295.2, 1465.3, 1705.4, 2850.9, 2919.4 cm-1; 1H-NMR (CDCl3) δ 0.9 (pent, 2H, -CH2), 1.20 (br, 15H, 5 × -CH3),  1.25 (br, 2H, -CH2), 1.63 (m, 2H, -CH2), 2.35 (m, 1H, -CH) ppm; 13C-NMR (CDCl3) δ 14.12 (-CH3), 22.69 (-CH-CH3), 24.70 (-CH2-CH3), 29.06 [-C (CH3)3], 29.24(-CH2-CH2-CH3), 29.70 (-CH2-(CH2)2-CH3), 31.93 [-CH (CH3) (CH2)], 195.56 (-C=O) ppm; MS (APCI) m/z 170 [M]+ (C11H22O), 172 [M+2]+, 141[M – CH2-CH3]+, 113  [M – (CH2)3-CH3]+, 57 [M – C=O-CH (CH3)-(CH2)3-CH3]+.

RESULTS AND DISCUSSION: Compounds 1-3 are being reported for the first time from S. media.

Compound 1 was isolated as white amorphous solid. The mass spectra of compound 1 showed [M]+ at m/z 170 suggesting the molecular formula C12H26 and [M+2]+ ion peak at m/z 172. The loss of 43 molecular units generated an ion at m/z 127 indicated the presence of [M - CH (CH3)2] +. Significant ions at m/z 85 and 57 correspond to [M – CH (CH3) (CH2)-CH (CH3)2] + and [M – CH (CH3)-CH (CH3)-CH2-CH (CH3)2] +. The IR spectrum featured bands at 722.9 cm-1 (long chain band), 1376.2 cm-1 (CH3 bending), and 1461.9 cm-1 (CH2 bending). Aliphatic C-H stretch featured at 2852.5 and 2920.8 cm-1. In the 1H NMR spectrum, signals appeared at δ 0.84 (br, 4H, 3-CH2 & 6-CH2), 0.86 (br, 2H, 2-CH & 7-CH), 1.25 (br, 18H, 1-CH3, 2′-CH3, 4′-CH3, 5′-CH3, 7′-CH3 & 8-CH3), 1.55 (m, 2H, 4-CH & 5-CH), ppm. In the 13C NMR spectrum, the signals showed at δ 14.09 (2C, 4′-CH3 & 5′-CH3), 22.67 (2C, 2′-CH3 & 7′-CH3), 29.68 (2C, 2-CH & 7-CH), 31.92 (2C, 4-CH & 5-CH), 37.10 (2C, 3-CH2 & 6-CH2) ppm. These data suggested the structure of this compound to be 2, 4, 5, 7-tetramethyloctane.

COMPOUND 1

Compound 2 was isolated as yellow gummy solid. The mass spectrum of compound 2 indicated an [M] + at m/z 216 suggesting the molecular formula C12H24O3 and [M+1] + at m/z 217. The loss of 43 molecular units generated an ion at m/z 173 indicated the presence of [M – CH (CH3)2] +. Significant ions at m/z 145, 131 and 103 correspond to [M – CH (CH3)2-(CH2)2] +, [M -CH (CH3)2-(CH2)3] + and [M – O-C=O-CH (CH3)-CH2OH]+ respectively. The IR spectrum showed bands at 1090.7 and 1157.4 cm-1 for C-O stretching of alcohol and ester respectively. OH stretch featured at 3426.0 cm-1 and C=O stretch featured at 1727.6 cm-1. In the 1H NMR spectrum, signals appeared at δ 0.80 (m, 2H, 5-CH2), 1.08 (br, 1H, 6-CH), 1.35 (br, 6H, 7-CH3 &   6′-CH3), 1.52 (m, 2H, 4-CH2), 1.72 (br, IH, 2′-CH), 1.83 (m, 2H, 3-CH2), 2.07 (m, 2H, 2-CH2), 2.35 (d, 3H, 2″-CH3), 2.70 (br, 1H, 3′-OH), 4.32 (t, 2H, 3′-CH2) ppm. Its 13C NMR signals appeared at δ 14.13 (1C, 2″-CH3), 22.70 (2C, 7-CH3 & 6′-CH3), 25.52 (1C, 3-CH2), 27.09 (1C, 4-CH2), 29.36 (1C, 6-CH), 29.66 (1C, 2-CH2), 37.09 (1C, 5-CH2), 50.04 (1C, 2′-CH), 57.36 (1C, 3′-CH2), 61.70 (1C, 1-CH2), 185.56 (1C, 1′>C=O) ppm. In light of these observations, the structure of 2 was elucidated as 6-methyl heptyl-3′-hydroxy-2′-methyl propanoate.

COMPOUND 2

Compound 3 was isolated as white crystals. The mass spectrum of compound 3 showed a molecular ion peak at m/z 170 calculated for C11H22O and [M+2] + at m/z 172. The loss of 113 molecular units from [M] + generated an ion at m/z 57 indicated the presence of tertiary carbon, correspond to [M – C=O-CH (CH3)-(CH2)3-CH3] +. Significant ions at m/z 141 and 113 indicated the presence of [M – CH2-CH3] + and [M – (CH2)3-CH3] +. The IR spectrum showed bands at 1295.2 and 1705.4 cm-1 for C-C=O-C bending and C=O stretching. In the 1H NMR spectrum, signals appeared at δ 0.9 (pent, 2H, 6-CH2), 1.20 (br, 15H, 1-CH3, 2′-CH3, 2″-CH3, 4′-CH3 & 8-CH3), 1.25 (br, 2H, 7-CH2), 1.63 (m, 2H, 5-CH2), 2.35 (m, 1H, 4-CH) ppm. In the 13C NMR spectrum, signals appeared at δ 14.12 (1C, 8-CH3), 22.69 (1C, 4′-CH3), 24.70 (1C, 7-CH2), 29.06 (3C, 2′-CH3, 2″-CH3 & 1-CH3), 29.24 (1C, 6-CH2), 29.70 (1C, 5-CH2), 31.93 (1C, 4-CH), 195.56 (1C, 3 >C=O) ppm. These data led to characterize compound 3 as 2, 2, 4-trimethyloctan-3-one.

COMPOUND 3

Studies on the constituents of aerial parts of Stellaria media Linn. have led to the isolation and characterization of three compounds ─ 2, 4, 5, 7-tetramethyloctane, 6-methylheptyl-3′-hydroxy-2′-methylpropanoate, 2,2,4-trimethyloctan-3-one.

CONCLUSION: The present study involved the isolation and characterization of three new compounds - Alkane (1) was separated as a white amorphous solid from petroleum ether extract of S. media. Bioactivity-guided fractionation of methanol extract of S. media ultimately led to the isolation of two compounds ― an ester (2) and a ketone (3). All the three compounds were reported for the first time in S. media Linn.

ACKNOWLEDGEMENT: This work was funded by the University Grants Commission (UGC).

CONFLICT OF INTEREST: Nil

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    How to cite this article:

    Arora D and Sharma A: Isolation and characterization of the chemical constituents of Stellaria media Linn. Int J Pharm Sci & Res 2014; 5(9): 3669-73. doi: 10.13040/IJPSR.0975-8232.5(9).3669-73.

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Article Information

13

3669-3673

388

1284

15

English

IJPSR

D. Arora and A. Sharma *

University Institute of Pharmaceutical Sciences, Panjab University, Chandigarh, India.

ans1959@rediffmail.com

28 February 2014

10 April 2014

24 May 2014

10.13040/IJPSR.0975-8232.5(9).3669-73

01 September 2014

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