Fatty Acids from Memecylon umbellatum (Burm.)
Himanshu Joshi1*, Arun B Joshi1, Hemlata Sati2, Gururaja MP1, Prajwal R Shetty1,
EVS Subrahmanyam1 and D Satyanaryana1
1Dept. of Pharma. Chem., NGSM institute of Pharmaceutical Sciences, Paneer, Deralakatte, Mangalore –574160 India.
2Department of Pharmacy, H.N.B Garhwal University. Srinagar Garhwal. Uttaranchal
*Corresponding Author E-mail: visitjoshi@yahoo.com.
ABSTRACT
The n-hexane extract of the roots of Memecylon umbellatum yielded, octocosonoic acid, cerotic acid, ethyl palmitate, palmitic acid and butyric acid. The structural elucidation of the isolated compounds was based on spectral data (IR, 1H NMR, 13C NMR, and mass spectroscopy).
KEY WORDS: Memecylon umbellatum, n-hexane extract, roots, fatty acids.
INTRODUCTION:
Memecylon umbellatum is a shrub or a small tree, belonging to the family Melastomataceae. It is found distributed in Western Peninsula, mostly on the coast. Inland mostly in evergreen and semi-evergreen shrub. It is also found distributed in Orissa, Assam, Sylhet, Tenasserim, Ceylon, Malay Peninsula-Malay Archipelago1,traditionally the plant is used as a cooling astringent; used in conjunctivitis as a lotion; and given internally in leucorrhoea and gonorrhea 2. The decoction of the root is used in excessive menstrual discharge. Apart from this various activities like anti diabetic3, anti viral4, and wound healing activity 5 have also been reported. From literature survey it has been revealed that various phytoconstituents such as include b-amyrin, sitosterol, oleanolic acid, ursolic acid, sitosterol-b-D-glucoside and umbelactone6 were isolated from the Arial parts of the plant. The present work describes the isolation and characterization of the active constituents from the n-hexane extract of the roots of Memecylon umbellatum.
MATERIAL AND METHODS:
General Methods:
All the melting points were recorded in a Toshniwal melting point apparatus and were uncorrected. IR spectra of the compounds were recorded using the KBr pellet method on a Nicolet Avator 330 FTIR, Perkin-
Elmer model 700 IR spectrophotometer. 1HNMR spectra of the compounds were taken on AMX 400 (270 MHZ) NMR spectrometer using CDCl3 as a solvent. Mass spectra were recorded on LCMS/GCMS. TLC was carried out using Silica-gel G (Merck).Column chromatography was carried out on Silica-gel (Merck, 70-230 mesh).
All the chemicals and reagents used were obtained in high purity either from S.D. fine chemicals Pvt. Ltd., Bombay, India or E-Merck Pvt. Ltd., Bombay, India.
Plant material:
Memecylon umbellatum roots were collected from Mangalore, Karnataka, India. during April 2006. The plant was aunthetified by Prof. Gopoal Krishna Bhat, department of Botany, Poornaprajna College, Udipi, Karnataka, India. A voucher specimen (No.102 a) was deposited in NGSM Institute of Pharmaceutical Sciences, Paneer, Mangalore, India.
Preparation of the ethanolic extract:
The roots were collected from Mangalore, Karnataka (India) during summer, washed, shade dried and then milled into a coarse powder by mechanical grinder. The powdered plant material (15 kg) was then soaked in ethanol and kept aside for four days. After four days the ethanolic layer was decanted off. The process was repeated for four times. The solvent from the total extract was distilled off and the concentrate was evaporated on a water bath to a syrupy consistency and then evaporated to dryness (1.560 kg).
Fractionation of ethanolic extract:
The extract (1.5 kg) was divided into five equal portions. Each portion was suspended in distilled water (500ml) and then extracted with n-hexane (8×500ml), petroleum ether (60-80˚C, 8×500ml), chloroform (8×500ml), butanol (8×500ml), and Ethyl acetate (8×500ml) in succession. All the fractions were then washed with distilled water (80ml), dried over anhydrous sodium sulphate and freed of solvent by distillation. The ethanolic extract was thus fractioned into n-hexane soluble extract (3.7g), petroleum ether (60-80˚C) soluble extract (52g), chloroform soluble extract (62 g), butanol soluble extract (92g), and ethyl acetate soluble extract (38 g).
n-hexane extract:
The residue (3 g) was dissolved in n-hexane (10 ml) and adsorbed onto silica gel (10 g). After evaporation of the solvent it was loaded onto a silica gel column (120 g) prepared in n-hexane. The column was eluted first with n-hexane, followed by n-hexane: ethyl acetate graded mixtures (95:5, 90:10, 80:20, 70:30, 60:40 and 50:50) and finally with 100% ethyl acetate. The elutions were monitored by TLC (Silica gel-G; visualization by Vanillin - Sulphuric acid reagent heated at 110ºC). Each time 5 ml were collected and identical elutes (TLC monitored) were combined and concentrated to 5 ml and kept in a refrigerator.
Elution’s carried out with n-hexane: ethyl acetate graded mixture (95:5) resulted a single spot on TLC. After removing the solvent, a sticky pale yellow residue resulted. It was designated as compound I (66 mg).Elution’s carried out with n-hexane, ethyl acetate graded mixture (90:10) resulted a mixture of two compounds, after removing the solvent a residue (130mg) resulted. This residue was subjected to preparative TLC in the solvent system n-hexane: ethyl acetate (90:10).The two compounds were thus isolated as pure compounds and designated as compound II (52 mg) and compound III (60 mg). Elution’s carried out with n-hexane:ethyl acetate graded mixture (80:20) resulted a single spot on TLC, after removing the solvent, resulted a white crystalline scales were obtained. The compound was designated as compound IV (59 mg).Elution’s carried out with n-hexane: ethyl acetate graded mixture (50:50) resulted a single spot, after removing the solvent, an oily colourless liquid resulted. The compound was designated as compound V (63mg).Elution’s carried out with 100% ethyl acetate gave a resinous mass which was not prossed further.
Octocosonoic acid (compound-I):
Sticky pale yellow, m.p. 83°C , IR νmax /cm-1: 3426.2, 2918.7, 2850,1726.6(C=O), 1657.1H NMR (400 MHz, CDCl3): δ 0.8696 (m, 3H, CH3), δ 1.2537 to δ 1.4066 (m, 48H, 24X CH2), δ 1.684 (s,2H , CH2 at C-3), δ 2.1743 (s,2H, CH2 at C-2).13CNMR(CDCl3): δ14.13 (CH3), δ174.2(COO), δ 29.71(CH2)22, δ 22.71 (CH2 α to CH3), 31.94 (CH2 β to CH3),δ 29.38 (CH2 γ to CH3), 34.28(CH2 α to CH3). LC-MS (m/z): 424(M+, C28H56O2,) the other peaks appeared at, 409 (100%) 397, 361, 307, 256, 191, 137, 87 and 66.
Cerotic acid (compound-II):
Green waxy semisolid, m.p.46°C. IR νmax /cm-1: 3382.3 cm-1 (br ,OH), 1702.55 cm-1 ( C=O str.), 1612.36 cm-1 (C=C str.), 1519.61 cm-1 (C-H deformation in CH3), 1038.15 cm-1 (C-H deformation in CH2). 1H NMR (400 MHz, CDCl3): δ13.98(CH3), δ178.71(COO), δ 29.14(CH2)21, δ 22.48 (CH2 α to CH3), 31.71 (CH2 β to CH3), δ 29.47 (CH2 γ to CH3). 13C NMR (CDCl3): δ13.98(CH3), δ178.71 (COO), δ 29.14(CH2)21, δ 22.48 (CH2 α to CH3), 31.71 (CH2 β to CH3),δ 29.47 (CH2 γ to CH3). LC-MS (m/z): 396 (M+, C4H8O2,) the other peaks appeared at 351, 338, 324, 310, 296, 281, 267, 253, 239, 225, 77, 40.
Ethyl palmitate (compound-III):
White wax, m.p. 42 °C, IR νmax /cm-1: 2917.4 cm-1 (C-H str. of CH3), 2847.9 cm-1 (C-H str. of CH2),1737.3 cm-1 ( C=O str.), 1655.3 cm-1 (C=C str.) 1H NMR (400 MHz, CDCl3) : δ 0.8626 to δ 0.8967 (m, 2×CH3, 6H, terminal methyl), δ 1.2536 to δ 1.300 (m, 26H,13×CH2), δ 2.2676 to δ 2.3054 (t, 1×CH2, 2H, at C-2), δ 4.0370 to δ 4.0706 (t, 2H, at ester linkage) 13C NMR (CDCl3): : δ14.13(CH3), δ173.33(COO), δ 29.71(CH2)11, δ 22.17 (CH2 α to CH3), δ 31.94 (CH2 β to CH3), δ 29.38 (CH2 γ to CH3), δ 66.10(ester linkage). 13C NMR (CDCl3): δ14.13(CH3), δ173.33(COO), δ 29.71(CH2)11, δ 22.17 (CH2 α to CH3), δ 31.94 (CH2 β to CH3),δ 29.38 (CH2 γ to CH3), δ 66.10(ester linkage). GC-MS (m/z): 284 (M+, C18H36O2,) the other peaks appeared at, 255, 239, 213, 215, 143, 101, 88, 70, 55, 44.
Palmitic acid (compound-IV):
White crystalline scales, m.p.63-64°C, IR νmax /cm-1: 3367.95 cm-1 (br ,OH),1702.95 cm-1 ( C=O str.), 1612.93 cm-1(C=C str.),1519.98 cm-1 (C-H deformation in CH3), 1038.90cm-1 (C-H deformation in CH2) 1H NMR (400 MHz, CDCl3) : δ 0.8432 to δ 0.9346 (m, 3H, terminal methyl),δ 1.2537 to δ 1.3329 (m, 13×CH2, 26H, of C-3), δ 2.1743 to δ 2.3054 (s, 1×CH2,2H, at C-2) 13C NMR (CDCl3): δ14.13 (CH3), δ173.70 (COO), δ 29.71(CH2)12, δ 22.71 (CH2 α to CH3), 31.94 (CH2 β to CH3). GC-MS (m/z): 256 (M+, C16H32O2,) the other peaks appeared at 192, 160, 128, 111, 85, 71, 57.
Butyric acid (compound-V):
Oily colourless liquid, m.p. 163.5°C, IR νmax /cm-1: 3398.3 cm-1 (br ,OH), 2925.5 cm-1 (C-H str. in CH3), 2853.8 cm-1 (C- H str. of CH2),1722.1 cm-1 ( C=O str.),1630.3 cm-1(C=C str.) 1H NMR (400 MHz, CDCl3) :δ 0.8626 to δ 0.8967 (m, 1×CH3, 3H, terminal methyl), δ 1.2536 to δ 1.300 (m, 1×CH2,2H, of C-3), δ 2.2676 to δ 2.3054 (t, 1×CH2, 2H, at C-2) 13C NMR (CDCl3):δ13.98 (CH3), δ178.02 (COO), δ 38.23 (CH2), δ19.14 (CH2 α to CH3). GC-MS (m/z): 88 (M+, C4H8O2) the other peaks appeared at 77, 57, 44, 40 (100%).
RESULTS AND DISCUSSION:
Octocosanoic acid 7:
CH3-(CH2)26-COOH, m.p. 81-81.2ºC (82ºC).The IR spectrum showed the characteristic absorption bands at 3426.2 cm-1, indicating the presence of hydroxyl group of carboxylic acid. The proton NMR signal at δ 0.86 indicated terminal methyl protons, δ 1.2-1.4 CH2 protons, δ1.68 CH2 of C-3 protons and δ 2.17 indicates CH2 protons adjacent to carboxylic group, respectively. Further signal at δ 174.2 (C-1) and δ 14.13 (C-28) in the 13C NMR spectra gave evidence for the above findings. The LCMS mass spectra showed the molecular ion peak at m/z 424[M]+ corresponding to molecular formula C28H56O2.
Cerotic acid8:
CH3-(CH2)24-COOH, m.p.46ºC .The IR spectrum showed the characteristic absorption bands at 3382.3 cm-1, indicating the presence of hydroxyl group of carboxylic acid. The proton NMR signal at δ 0.86 indicated terminal methyl protons, δ 1.2-1.3 CH2 protons, δ 2.26 indicates CH2 protons adjacent to carboxylic group, respectively. Further signal at δ 178.7 (C-1) and δ 13.98 (C-26) in the 13C NMR spectra gave evidence for the above findings. The GCMS mass spectra showed the molecular ion peak at m/z 396[M]+ corresponding to the molecular formula C26H52O2.
Ethyl palmitate7:
CH3-(CH2)14-COO-C2H5 m.p. 42ºC .The IR spectrum showed the characteristic absorption bands at 1702.55 cm-1, indicating the presence of carbonyl group, and absence of hydroxyl group respectively. The proton NMR signal at δ 0.86 indicated terminal methyl protons, δ 1.2-1.3 CH2 protons, δ 4.03 CH2 protons, and δ 2.26 indicates CH2 protons adjacent to carboxylic group, respectively. Further signal at δ 173.33 (C-1) and δ 66.10 (ester linkage) in the 13C NMR spectra gave evidence for the above findings. The GCMS mass spectra showed the molecular ion peak at m/z 284[M]+ corresponding to molecular formula C18H36O2.
Palmitic acid8,9:
CH3-(CH2)14-COOH m.p. 63-64ºC. The IR spectrum showed the characteristic absorption bands at 3367.9 cm-1, indicating the presence of hydroxyl group of carboxylic acid. The proton NMR signal at δ 0.84 indicated terminal methyl protons, δ 1.2-1.3 CH2 protons, and δ 2.17 indicates CH2 protons adjacent to carboxylic group, respectively. Further signal at δ 173.7 (C-1) and δ 14.13 (C-16) in the 13C NMR spectra gave evidence for the above findings. The GCMS mass spectra showed the molecular ion peak at m/z 256[M]+ corresponding to molecular formula C16H32O2.
Butyric acid10:
CH3-(CH2)2-COOH, b.p. 163.5ºC .The IR spectrum showed the characteristic absorption bands at 3398.3 cm-1, indicating the presence of hydroxyl group of carboxylic acid. The proton NMR signal at δ 0.86 indicated terminal methyl protons, δ 1.2-1.3 CH2 protons, and δ 2.17 indicates CH2 protons adjacent to carboxylic group, respectively. Further signal at δ 178.02 (C-1) and δ 13.98 (C-4) in the 13C NMR spectra gave evidence for the above findings. The GCMS mass spectra showed the molecular ion peak at m/z 88[M]+ corresponding to molecular formula C4H8O2.
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Received on 18.03.2009 Modified on 22.05.2009
Accepted on 26.05.2009 © AJRC All right reserved
Asian J. Research Chem. 2(2): April.-June, 2009 page 178-180