Qualitative and Quantitative Phytochemical Screening of Cleome Viscosa Linn. (Cleomaceae) leaves, Root and stem extract

All angiospermic plants are confined with treasure house of potential drugs. Drugs from plants are easily available, less expensive, safe, efficient and have no side effects. (Dewick, PM, 1996)). The curative properties of medicinal plants are mainly due to the presence of various complex chemical compounds in varied composition which occur as secondary metabolites. (Trease, GE et al., 1989). The existing secondary metabolites form the backbone of modern medicine (Karthikeyan et al., 2009). The phytochemical research based on ethnopharmacology is considered an effective approach in the finding of new anti-infective agents from higher plants (Edeoga, HO et al., 2005). In the present study, the prime focus is to evaluate the phytochemical constituents present in root, stem and leaves of Cleome viscosa Linn. (CLEOMACEAE).

• Test tube, Beaker, Conical flask.

• Ethanol

• Methanol

• Whatman filter paper no. 1

• Centrifuge

• Chloroform

• Distilled water

• Digital balance

• Incubator

• Water bath

• FeCl3

• Measuring cylinder 10ml capacity

• Extraction of plant materials : About 100 mg of dried and powdered plant materials was kept overnight in 25 ml of different solvents namely aqueous, ethanol and methanol. The extracts were filtered using whatman filter paper.

 Qualitative evaluation of phytochemical : The extracts were subjected to preliminary qualitative phytochemical screening following standard methods (Mali RG, 2010; Harborne, JB, 1998) 1. Test for alkaloids: 2 ml filtrate was taken in a clean test tube and added 1% HCL. The mixture solution was changed to cream colour which confirmed the test of alkaloid. 2. Test for cardiac glycosides: 2 ml filtrate was taken in a clean test tube and added 1 ml glacial acetic acid, FeCl3 and conc H2SO4. A brown ring confirmed the test of cardiac glycosides. 3. Test for phenols : 2 ml filtrate was taken in a clean test tube and added 1 ml of 1% FeCl3. Brown haziness appeared which confirmed the test of phenols. 4. Test for saponins: 1 ml filtrate was mixed with 5 ml distilled water and shaken vigorously for getting a stable froth. The persistence of frothing confirmed the test of saponin. 5. Test for steroids: 2 ml of filtrate was taken in a clean test tube and added 2 ml acetic analydride and conc. H2SO4. Blue-green ring confirmed the test of steroids.

reddish-brown colouration at the interface was formed which showed the +ve result of the presence of terpenoids. The results were presented in table-1.

+ = Detected ++ = Detected in higher amount - = Not detected Quantitative evaluation of phytochemical: 1. Quantification of total alkaloids: Total alkaloid content was estimated by following the method reported by Singh et al., (2004). For estimation of alkaloids, 100 mg powdered plant sample was placed in 10 ml of 80% ethanol overnight and filtered by whatman filter paper and then centrifuged at 5000 rpm for 10 minutes. The supernatant obtained was used for the estimation of total alkaloids. The reaction mixture contained 1 ml plant extract + 1 ml of 0.025 M FeCl3 in 0.5 M HCl and 1 ml of 0.05 M of 1, 10-Phenanthroline in ethanol. The mixture was warmed in hot water bath. The absorbance of red colour complex was measured at 510 nm against blank. Alkaloid content was estimated using the standard curve of colchicines. 2. Quantification of total cardiac glycosides: The plant sample was suspended in 25 ml distilled water. Then 2 ml conc. H2SO4 was added to it and was soon refluxed for 6-8 hours, cooled and extracted with chloroform

% of cardiac glucoside = where, B = Weight of beaker with extract A = Weight of empty beaker 3. Quantification of total saponins: 1 gm sample was taken into a conical flask and added 25 ml 20% methanol. The mixture solution was warmed in water bath for 4 hours at 35OC. The mixture solution was filtered and the residue was re-extracted with another 25 ml of 20% methanol. The combined extracts were reduced to 15 ml over a water bath at 90OC. The concentrate was then transferred to a separating funnel and 20 ml of diethyl ether was added and shaken vigorously. The aqueous layer was retained while the ether layer was discarded. The aqueous layer was further separated by 60 ml (2ml x 30 ml) of butanol. The combined butanol layer was washed with 10 ml of 5% NaCl twice. The extract was then transferred to a pre-weighed beaker and dried in a oven to a constant weight. The weight of the extract was the saponin content. % of saponins = x 100 4. Quantification of total phenols: 0.5 gm powdered sample was taken in a mortar and grinded with pestle and added 5 ml 80% methanol. The homogenate was centrifuged at 1000 rpm for 20 minutes. The supernatant was evaporated to dryness. The residue was dissolved in 20 ml distilled water. A volume of 0.2 ml was taken in a clean test tube and was made 3 ml by adding distilled water. Again 0.5 ml Folin-ciacaten reagent was added. Just after 3 minutes, 2 ml of 20% NaCl was added. The reaction mixture was then placed in water bath for 1 minute. Cooled and the absorbance was taken at 650 nm against the blank. The phenolic content was calculated by applying following formula – % phenol = 10 0 RESULT AND DISCUSSION The qualitative phytochemical profile of root, stem and leaves of Cleome viscosa is provided in table-1. The result showed the presence of alkaloids in all the extracts except methanolic extract of leaves. Cardiac glycosides were detected in ethanolic and methanolic extracts of all plant parts but not in aqueous extracts. Steroids were not detected in any

0

0.5

1

1.5

RootStemLeaves

Alkaloids% Cardiac Glycosides % Phenol

Saponins

(0.051%), Phenol (0.372%) and saponins (0.073%), Preliminary phytochemical screening actually helps in isolating and characterizing the chemical constituents present in the plant extracts which is necessary to understand herbal drugs and their preparations.

1. Dewick, PM, 1996. Tumor inhibitor from plants. Philadelphia. 2. Trease, GE, Evans, WC, 1989. A textbook of pharmacognosy. 3. Karthikeyan, AK, Shanthi, V., Nagasthaya, A., 2009. Priminary phytochemical and antibacterial screening of leaf extract of Adhatoda vasica. Int, Journ. Green Pharm. 3 : 78-80. 4. Edeoga,, HO, Okwu, De, Mbaebie, BO, 2005. Phytochemical constituents of medicinal plants. Afr. Journ. Biotechnol. 4 : 685-88. 5. Mali, RG, 2010. Cleome viscosa : A review on phytochemistry and pharmacology. Pharm. Biol. 48: 105-12. 6. Harborne, JB, 1998. Phytochemical methods, 3rd ed London. PP 1-302. 7. Singh, DK, Srivastava, B., Sahu, A. 2004. Spectrophotometric determination of Ranwolfia alkaloids. Anal Sc. 20 : 571-73. 6.

Research Scholar, Dept. Biotechnology, M.U.Bodhgaya, Bihar