preliminary phytochemical screening of...

ABSTRACT :In the present study the aqueous, methanol, petroleum ether, acetone and ethyl alcohol extract of the aerial part of Leonotis nepetaefolia was screened for the phytochemical analysis The extract showed the presence of phenolics, Alkaloids, saponin and phytosterol, carbohydrates, oils and fats, saponin, proteins and amino acids tannins, gums and mucilage, flavonoids and cardiac glycosides in different plant parts like stem, leaves and inflorescence. All the plant parts were found to contain alkaloids except in inflorescence ethyl alcohol and acetone extract it is absent. Carbohydrates present in all the extracts of all the three parts of the plant studied. Stem extract showed absence of fixed oils and fats in acetone extract, leaf extract in petroleum ether and aqueous extract and inflorescence for ethyl alcohol and aqueous extract respectively. Saponin present in all the extracts except petroleum ether, acetone and methanolic extract of inflorescence. Phenolic compound present only in leaf and stem extract. In view of importance of plants as remedy to diseases, there is an urgent need for a proper and systematic investigation of plants to isolate pharmacologically active ingredients.

Key words : Phytochemistry, crude extracts

INTRODUCTION:

PRELIMINARY PHYTOCHEMICAL SCREENING OF LEONOTIS NEPETAEFOLIA, AN AYURVEDIC HERB

U. G. BASARKAR* AND V. SHINDEP. G. Department of Botany

G. E. Society's, HPT Arts RYK Science College, Nashik 422 [email protected]

There is ample literature on preliminary phytochemical surveys and the knowledge of chemical constituents of plants is desirable to understand herbal drugs and their preparations. Most importantly, these studies will be helpful to isolate and characterize the chemical constituents present in those plant extracts. In addition, the knowledge of chemical constituents of plants would further be valuable in discovering the actual value of folkloric remedies (Farnsworth, 1966).

Labiatae family is largest and widely spread in Nasik district. It is almost cosmopolitan family of about 220 genera and 3500 species distributed all over the world (Curitus, 1881). In Nasik district about 15 genera and 59 species are found. Members of this family are mostly aromatic herbs or shrubs characteristically bearing essential oils.

Leonotis nepetaefolia belonging to family Labiatae is known to be native to tropical Africa and southern India. Whole plant is medicinally important especially roots of the plant have been used in Brahat Guduchi Taila in Ayurvedic formulations and also used in indications of Swasa (Asthma and Bronchitis), Kandu (Fever) and Visa (Poisonous conditions) (Anonymous, 2003). Seeds are used in burns in India (Kala, 2005). Leaves have been used in diabetes in Trinidad. Therefore by considering immense medicinal importance of the plant the present paper reports some phytochemical and physicochemical properties of different solvent extracts of leaves, stem and inflorescence of Leonotis nepetifolia an ayurvedic herb.

MATERIALS AND METHODS :

a) Collection , identification and processing of plant material : Fresh plant material was collected from the Nasik and nearby areas of Nasik. Plant was correctly identified with the help of Flora of Maharashtra and Flora of Nasik district. Plant material washed under running tap water and dried in the sun light. It was then homogenized to fine powder with electric

blender and stored in airtight bottles. This sample was used for extraction of organic compound.

b)Extraction of organic crude material from Leonotis

nepetaefolia : The 50 gm sample (leaves, stem and

inflorescence) weighed separately and used for soxhlation. SOLVENT USED : Depending on polarity the following solvents are selected1)Distilled water 2) Ethyl alcohol 3) Acetone 4) Petroleum ether 5) Methanol

c)Phytochemical analysis of plant extract : The

phytochemical are essential to metabolism and chemical

processes of plant body. The phytochemical are studied are

alkaloids terpenoids & steroids, flavonoids, glycosides,

tannins & saponin.IDENTIFICATION TEST : The test were done to find the presence of active chemical such as alkaloids , glycosides, terpenoids, steroid, flavonoids ,saponin, tannin by the following procedureALKALOIDSDetection of Alkaloids (Evans, 2002) : Solvent free extract, 50 mg is stirred with few ml of dilute hydrochloric Acid and filtered. The filtered is tested carefully with various alkaloid reagents as follows:A. Mayer's test : To a few ml of filtrate, a drop or two of Mayer's reagent are added by the side of the test tube. A white or creamy precipitate indicates the test as positive.Mayer's Reagent : Mercuric chloride (1.358 g) is dissolved in 60 ml of water and potassium iodide (5.0 g) is dissolved in 10 ml of water. The two solutions are mixed and up to 100 ml with water.B. Wagner's (Wagner, 2004) : To a few ml of filtrate, few drops of Wagner's reagent are added by the side of the test tube. A reddish- brown precipitate confirms the test as positive.Detection of Carbohydrates and Glycosides : The extract (100 g) is dissolved in 5 ml of water and filtered. The filtered is subjected to the following tests.a) Mayer's test : To 2 ml of filtered, two drops of alcoholic solution of -naphthol are added, the mixture is shaken

BIONANO FRONTIER VOL. 5 (2 - II) NOV. 2012ISSN 0974-0678239

well and 1 ml of concentrated sulphuric acid is added slowly along the sided of the test tube and allowed to stand. A violet ring indicates the presence of carbohydrates.b) Barfoed's test : To 1 ml of filtered, 1 ml of Barfoed's reagent is added and heated on a boiling water bath for 2 min. Red precipitate indicates presence of sugar.Barfoed's reagentCopper acetate, 30.5 g is dissolved in 1.8 mL of glacial acid.c) Benedict's test : To 0.5 ml of filtrate, 0.5 ml of Benedict's reagent is added. The mixture is heated on a boiling water bath for 2 min. A characteristic colored precipitate indicates the presence of sugar.d) To 3 ml of the aqueous extract was added about 1 ml of Iodine solution. A purple color at the interphase indicates the presence of carbohydrates.e) Keller Kiliani test 2 ml of extract was dissolved in 2 ml of glacial acetic acid containing one drop of ferric chloride solution. The mixture was then poured into the test tube containing 1 ml of concentrated sulphuric acid. A brown ring at the interphase indicates the presence of deoxy sugar, characteristics of cardenolides.Detection of Saponin :The extract (50 mg) is diluted with distilled water and made up to 20 ml. The suspension is shaken in a graduated cylinder for 15 min. A two cm layer of foam indicates the presence of saponin.Detection of Proteins and Amino acids :The extract (100 mg) is dissolved in 10 ml of distilled water and filtered through Whitman No.1 filter paper and the filtrate is subjected to tests for proteins and amino acids.

a) Million's test : To 2 ml of filtrate, few drops of Million's reagent are added. A white precipitate indicates the presence of proteins.b) Ninhydrin test : Two drops of ninhydrin solution (10 mg of ninhydrin in 200 ml 0f acetone) are added to two ml of aqueous filtrate. A characteristic purple color indicates the presence of amino acids.Detection of Phenolic compounds and Tanninsa) Ferric chloride test : The extract (50 mg) is dissolved in 5 mL of distilled water. To this, few drops of neutral 5% ferric chloride solution are added. A dark green color indicates the presence of phenolic compounds.Detection of Gum and Mucilage : The extract (100 mg) is dissolved in 10 ml of distilled water and to this 25 ml of absolute alcohol is added with constant stirring. White or cloudy precipitate indicates the presence of gums and mucilage.Glycoside :Glycosides are compounds which upon hydrolysis give rise to one or more sugars (glycones) and a compound which is not a sugar (aglycone or genine). To the solution of the extract in glacial acetic acid, few drops of ferric chloride and concentrated sulphuric acid and observed for a reddish brown coloration at the junction of two layers and the bluish green color in the upper layer. Terpenoid and steroids : Four milligrams of extract was treated with 0.5 ml of acetic anhydride and 0.5 ml of chloroform. Then concentrated solution of sulphuric acid was added slowly and red violet color for steroids (Siddiqui and Ali, 1997)Flavonoids :Four ml of extract solution was treated with 1.5

ml of 50% methanol solution. The solution was warmed and metal magnesium was added. To this solution, 5 - 6 drops of concentrated hydrochloric acid was added and red color for flavones.Tannins :To 0.5 ml of extract solution 1 ml of water and 1-2 drops of ferric chloride solution was added. Blue color was observed for Gallic tannins and green black for catecholic tannins. Fixed oils and fats : a) A small quantity of extract is pressed between two filter papers. Oil stain on the paper indicates the presence of fixed oil.b) Saponification test A few drops of 0.5 N alcoholic potassium hydroxide solutions are added to a small quantity of extract along with a drop of phenolphthalein. The mixture is heated on water bath for two hours. Formation of soap or partial neutralization of alkali indicates the presence of fixed oils and fats.

RESULTS AND DISCUSSIONS :

Plant material like leaves, stem and inflorescence was subjected to successive extraction with distilled water, ethyl alcohol, acetone, petroleum ether and methanol. Results of physicochemical properties are shown in table 1, 2 and 3. Phytochemical studies of different crude extracts revealed presence of alkaloids, carbohydrates, steroids, saponin, tannins, and phenols etc (Table 4, 5 and 6). All the plant parts were found to contain alkaloids except ethyl alcohol and acetone extract of inflorescence. Carbohydrates present in all the extracts of all the three parts of the plant studied. Stem extract showed absence of fixed oils and fats in acetone extract, leaf extract in petroleum ether and aqueous extract and inflorescence for ethyl alcohol and aqueous extract respectively. Saponin present in all the extracts except petroleum ether, acetone and methanolic extract of inflorescence. Phenolic compound present only in leaf and stem extract.

In 1997 Hirazawa et al selected six species of the family Labiatae as plant drugs to treat type I allergic diseases in traditional Sino-Japanese medicine. the main constituents in these plants likely to be effective for allergic diseases were flavonoids, volatile oils, tannins, sterols, phenols, saponin and many kinds of alkaloids. According to Hirazawa, these constituents probably act by inhibiting the histamine release from mast cells, direct antihistaminic reaction, anti-inflammatory actions. The results revealed the presence of medicinally active constituents in the present plant studied. The phytochemical compounds identified in this study have earlier been proved to be bioactive. The presence of some of these compounds have been confirmed by previous workers to have medicinal as well as physiological activity and therefore could be said to be responsible for the efficacy of the leaves of the plants studied in treatment of different ailments. The plant extracts could therefore be seen as a potential source for useful drug. The continued traditional medicinal use of this plant is therefore encouraged while it is suggested that further work should be carried out to isolate, purify and possibly characterize the active constituents responsible for the activity of this plant. Also additional work should be embarked upon with a view to elucidate the possible mechanism of action of these extracts.


Table 1 physicochemical characteristic of different extracts of Leonotis nepetaefolia leaf

Table 2 physicochemical characteristic of different extracts of Leonotis nepetaefolia stem

Table 3 physicochemical characteristic of different extracts of Leonotis nepetaefolia inflorescence

REFERENCES :


1.Anonymous, Ayurvedic Formulary of India, Part I, (Government of India, Ministry of Helth and Family wefare, Department of ISM and H), (2003), 75.

2. Curitus, T. (1881),J. Prak. Chem. , 34, 239.

th3. Evans WS (2002) Trease and Evan S Pharmacognosy. 5 ed, Haaarcourt brace ad company, pp, 336

4. Hizarawa, M., Higashino, H. and Komai, K. (1997) Kinki Daigakh Nogakubu Kiyo, 30, 31.

5. Kala, C. P., Ethanomedicinal Botany of the Apatani in the Eastern Himalayan regions of India, J. Ethnobiol Ethnomed, 1 (2005), 11.

6.Wagner, H. and Bladt, S., Plant Drug Analysis, A Thin Layer nd stChromatography Atlas, 2 edn and 1 Indian Reprint, (Springer Pvt.,

Ltd., New Delhi, India), 2004.

Table 4 Preliminary phytochemical analysis of crude extract of Leonotis nepetaefolia Stem

(+) minimum presence, (++) maximum presence, (-) absent

Table 5 Preliminary phytochemical analysis of crude extract of Leonotis nepetaefolia Leaf




Table 6 Preliminary phytochemical analysis of crude extract of Leonotis nepetaefolia Inflorescence


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