Isolation and Structure Elucidation of Four Antioxidant Compounds from Sudanese Sonchus oleraceus Plant

Abstract

Sonchus oleraceus (moleita) from the family Asteracae was investigated for its radical scavenging activity. The whole herb was firstly extracted with dicloromethane to remove as much of chlorophyll and other colouring pigments. The plant residue was extracted with methanol. The methanol extract was tested for its antioxidative contents. The methanolic extract showed 88% radical scavenging activity against DPPH(1,1-Diphenyl-2-picyl hydrazyl) a reagent used to test compound for their ability to neutralize free radicals. The methanolic extract of the whole plant was dissolved in 500ml distilled water. The mixture was poured into a separatory funnel and successive addition of 200 milletres of ethyl acetate was added with continuous shaking. Extracts of the ethyl acetate layer were collected. The ethyl acetate was chosen as a solvent that is optimum for extracting antioxidant (polyphenolic) compounds. Primary screening of the ethyl acetate extract showed saponins, coumarins, tannins, alkaloids, flavonoids, anthraquinones, sterols, triterpines and phenols. The highest concentrations were for tannins, flavonoids and phenols. The ethyl acetate layer was isolated into fractions with column chromatography. Solvent eluent mixtures were used from least to highest polarities (petroleum ether, chloroform, ethyl acetate, and methanol). 14 fractions showed high antioxidant activity. (Fraction 23 –Fraction 36).Fractions were monitored with TLC (Thin Layer Chromatography) and suitable solvent systems. Fraction 23 gave 69% activity and 1.99 gm yield and solvent system in the ratio (4 : 6) chloroform ; ethyl acetate. Fraction 23 was purified with IV preparative thin layer chromatography to give three compounds. Solvent system for purification was (toluene: ethyl acetate:formic acid) 5:4:1. Fraction 23 purified to give (23_2 and 23_3) which were analysed for structure characterization.Structure chrachterization was done with different spectroscopic techniques:Infra-red, Mass, and Nuclear Magnetic Resonance (NMR). Fraction 24showed 88% RSA (radical scavenging activity) when analysed with DPPH.Weight of fraction was 0.640 gm.Solvent system for separation was chloroform: ethyl:acetate (3:7). Solvent system for purification was (toluene: ethyl.acetate: formic acid) 5:4:2.Fraction 24 gave one compound for structure characterization . Sub-column was carried for fractions (30+31) using a smaller column. The solvents used were only ethyl acetate and methanol. Nine sub-fractions were isolated. Sub fraction 7 was chosen for the fourth compound. It gave 40% RSA against DPPH, but its separation was good on TLC .Solvent system for isolation was Ethyl acetate: methanol (4:6). Four compounds were isolated, compound (1) Lignan glycoside. Brown – green solid, with a solvent system of isolation Toluene: Ethyl Acetate: Formic Acid in the ratio 5: 4: 2. The molecular ion was m/z 869. Weighting 0.466 gm. Compound (2) Chalcone glycoside. Yellow-Brown solid. Solvent system of isolation (Toluene: Eth. Acet. Form.acid) in the ratio 5:6:3.The molecular ion was m/z 745. Weighting 0.121 gm. Compound (3) Flavone glycoside. Yellow- brown solid. Solvent system for isolation (Tol: Eth.Acet : Formic Acid). The molecular ion m/z 745.Weighting 0.530 gm. V Compound (4)Tannic acid. Dark brown amorphous solid. Solvent system for isolation was Ethyl Acetate: Methanol (4:6). m/z 789 Weight 0.11 gm. This investigation proved that Sonchus oleraceus plant possesses high activity against RSA, which is prominent for protection of cells against harmful radicals.