INTRODUCTION:

The plant kingdom is a treasure house of potential drugs and in the recent years there has been an increasing awareness about the importance of medicinal plants. Drugs from the plants are easily available, less expensive, safe and efficient and rarely have side effects. Plant extracts and compounds derived from plants are in use as drug from the ancient times (Ahmed John and Koperuncholan, 2012).

In India, the practice of using plants for treating a wide variety of diseases is being carried out over ages and this practice is well explained and studied in Ayurveda (Siddharthan, 2007). Over one and half million practitioners of Indian system of medicine in the oral and codified streams use medicinal plants in preventive, promotive and curative applications (Madhulika, 2010). The booming of traditional medicine industry is increasing demand on medicinal plant products and 90% of the medicinal plants come from natural habitats (Raju, 2007). During the past few decades enormous efforts were undertaken to introduce new chemical entities with potential medical applications in drug discovery research (Grover, 2002; Marcy, 2005).

Herbalism ("herbology" or "herbal medicine") is use of plants for medicinal purposes, and the study of such use. Plants have been the basis for medical treatments through much of human history, and such traditional medicine is still widely practiced today. The demand for plant based medicines, health products, pharmaceuticals, food supplement, cosmetics etc are increasing in both developing and developed countries, due to the growing recognition that the natural products are non-toxic, have less side effects and easily available at affordable prices (Evans, 1994). The use of herbal medicine is becoming popular due to toxicity and side effects of allopathic medicines. This led to sudden increase in the number of herbal drug manufactures (Agarwal, 2005).

Phytochemistry is a distinct discipline somewhere in between organic chemistry, plant biochemistry and closely related to natural products. It deals with a variety of organic substances accumulated in plants. The plant may be considered as a biosynthetic laboratory. The qualitative and quantitative estimation of the phytochemical constituents of a medicinal plant is considered to be an important step in medicinal plant research (Kokate, 1994).

Phytochemicals are bioactive chemicals of plant origin. They are naturally synthesized in all parts of the plant body, bark, leaves, stem, root, flower, fruits, seeds etc (ie) any part of the plant body contain active components (Criagg and David, 2001; Tiwari et al., 2011). These chemicals are often referred to as “secondary metabolites” of which there are several classes including alkaloids, flavanoids, coumarins, glycosides, gums, polysaccharides, phenols, tannins, terpenes and terpenoids (Harborne, 1973; Okwu, 2004). The most important and commonly encountered secondary metabolites of plants are saponins, tannins, flavanoids, alkaloids, anthraquinones, cyanogenic glycosides, carbohydrates, terpenoids and flavanoids (Edoga et al., 2005). The presence of these secondary metabolites in plants probably explains the various uses of plants for traditional medicine. Knowledge of the chemical constituents of plants is desirable, not only for the discovery of therapeutic agents but also because such information may be of value in disclosing new sources (Lena, 2010).

Medicinal drugs can sometimes be better than using medical drugs. Because, the herbal medicines are easily available, relatively cheaper cost and non-toxic nature when compared to modern medicine (Uniyal, 2006). Herbal medicines also reduce the side effects and effectives with chronic conditions. If herbs are used correctly, they will help to treat a variety of conditions and in some cases, may have fewer side effects than some conventional medications (Calixto, 2000).Herbs relieve the symptoms of asthma (Grover et al., 2002). Many herbal antibiotics have direct germ killing effects, they have as a primary action, the stimulation of the body’s own immune response. Herbs that can be applied to the skin to prevent the growth of bacteria. Cooling herbs used to reduce or prevent fever. It also calm nervous tension and nourish the nervous system (Mishra et al., 2000).

Helminthes infections are commonly found in community and being recognized as cause of much acute as well as chronic illness among the various human beings as well as cattle’s. More than half of the population of the world suffers from various types of infection and majority of cattle’s suffers from worm infections (Chaturvedi et al., 2009). However, the high cost of modern anthelmintics has limited the effective control of these parasites. In some cases widespread intensive use of sometimes low quality anthelmintics (Monteiro et al., 1997) has led to development of resistance and hence a reduction in the usefulness of available anthelmintics (Waller, 1997). Although the use of alternate drugs has also been advocated as a measure to avoid the development of resistant strains of helminth parasites and as a means of reducing the cost of controlling helminthic diseases (Kelly and Hall, 1979; Okon et al., 1980; Taylor and Hunt, 1989; Coles and Roush, 1992).The plants are known to provide a rich source of botanical anthelmintics (Geert and Dorny, 1995; Coles, 1997; Satyavati et al., 1976). A number of medicinal plants have been used to treat parasitic infections in man and animals (Akhthar et al., 2000). Anthelmintics derived from plants can be a solution to this world wide problem as they form safe and non-toxic agents with an altered site of action (Maciel et al., 2006; Akhthar et al., 2000).

Adhatoda vasica

Adhatoda vasica, commonly known as malabar nut, adulsa, adhatoda, vasa, or vasaka, (Aslam et al., 2013) is a medicinal plant native to Asia, widely used in Siddha Medicine, Ayurvedic and Unani systems of medicine. It is part of the Acanthaceae plant family. It is a perennial, evergreen and highly branched shrub (1.0 m to 2.5 mm height) with bitter taste. It has opposite ascending branches with white, pink or purple flowers (Patel and Venkata-Krishna- Bhatt, 1984). When the leaves are dried, they appear dull brownish green in color and taste bitter. The corolla is large and white, with lower lip streaked purple. The stomata in the plant are elongated and oval in shape.

In Ayurvedic medicine, malabar nut (Adhatoda vasica) has been used for a multitude of disorders including; bronchitis, leprosy, blood disorders, heart troubles, thirst, asthma, fever, vomiting, loss of memory, leucoderma, jaundice, tumors, mouth troubles, sore-eye, fever, and gonorrhea. A decoction of the leaves of Vasaka may be used to help with cough and other symptoms of colds. A poultice of the leaves of Vasaka may be applied to wounds for their antibacterial and anti-inflammatory properties. The poultice is also helpful in relieving rheumatic symptoms when applied to joints. Vasaka has been used to control both internal and external bleeding such as peptic ulcers, piles and bleeding gums. Vasaka exhibits antispasmodic, expectorant and blood purifying qualities.

Materials and Methods:

Selection of Plant Material:

Adhatoda vasica, commonly known as Malabar nut was one of the member of the family Acanthaceae. Adhatoda vasica was selected for the present study.

Plate 1: Adhatoda vasica

Preparation of powder from Plant Parts:

The plant parts such as root, leaf and stem of Adhatoda vasica were collected freshly and transported to the laboratory, they were then washed thoroughly in running tape water and then with distilled water. The whole plant parts were cut into small bits to facilitate shade drying and the drying process was continued to decrease the moisture content. After drying, the plant materials were ground well using mechanical blender into fine powder. Then the powder was stored in airtight containers with proper labeling and kept in refrigerator for further use.

Preparation of plant extract: (Percolation process):

For the percolation process, the macerated plant powders were soaked in solvents such as Ethanol, Ethyl acetate, Chloroform, Aqueous and Hexane individually. Extraction was done by soaking one part of plant powder to three parts of liquid solvent (1:3) and kept for percolation process for 3-5 days. Then the crude extracts were filtered using Whatman No.1 filter paper, evaporated and concentrated into solid extracts under room temperature.

Phytochemical analysis:

The extracts of each solvent was used to analyse the presence of different phytochemical constituents. The phytochemical analysis was done using the standard methods (Harborne, 1973; Trease and Evans, 1989; Sofowara, 1993; Harborne, 2005; Kumar et al., 2009).

Test for Quinones:

To 1 ml of extract, 1 ml of Conc.H₂SO₄ was added. Formation of red colour indicates the presence of quinones.

Test for Coumarins:

A few drops of ammonia were added on a filter paper. To this, a drop of the extract was added and the paper was observed for fluorescence.

Test for Flavanoids:

Ferric Chloride Test:

The extract was treated with a few drops of FeCl₃ solution. Formation of a blackish red colour indicates the presence of flavanoids.

Test for Gums and Mucilages:

About 5 ml of the extract was slowly added to 5 ml of absolute alcohol under constant stirring. The appearance of precipitation indicates the presence of gums and mucilages.

Test for Tannins:

To 1 ml of the solvent extract, few drops of 1% FeCl₃ solution was added. The appearance of a blue, black, green or blue green precipitate indicates the presence of tannins.

Test for Resins

Acetone-H₂O Test:

The extracts were treated with acetone. A small amount of water was then added and shaken. Appearance of turbidity indicates the presence of resins.

Test for Terpenoids:

Salkowski Test:

To 1 ml of the solvent extract, 2 ml of chloroform was added. Then 3 ml of Conc.H₂SO₄was added carefully to form a layer. A reddish brown coloration of the interface indicates the presence of terpenoids.

Test for Phenols:

Ferric Chloride Test:

To 1 ml of solvent extracts, 3 ml of distilled H₂O was added. To this, a few drops of neutral 5% FeCl₃ solution was added. Formation of a dark green colour indicates the presence of phenolics.

Test for Saponins:

Foam Test:

About 2 ml of distilled H₂O and 1 ml of solvent extract were mixed and shaken vigorously. Formation of a stable persistent froth indicates the presence of saponins.

Test for volatile oils:

To 1 ml of the extract, 1 ml of 90% ethanol was added, followed by the addition of a few drops of FeCl₃ solution. Formation of a green colour indicates the presence of volatile oils in the given sample.

Anthelmintic activity:

Test drug:

The leaves of Adhatoda vasica was cut into small pieces dried in shade and powdered coarsely. The powdered leaves of Adhatoda vasica was extracted exhaustively with increasing polarity solvents (hexane, chloroform, ethyl acetate, ethanol, and aqueous) for 72 hours followed by 48 hours and 24 hours.

Reference drug:

Albendazole was prepared by dissolving them in normal saline at a concentration of 15mg/ml.

Worm collection:

Indian earthworm Pheretima pothuma (Annelida) were collected from the waterlogged areas of soil. The average size of Pheretima pothuma was 6-8 cm long. They were washed with water to remove dirt.

Procedure:

Indian earth worm Pheretima pothuma has anatomical and physiological resemblance with the intestinal round worm parasites of human beings. Pheretima pothuma was placed in petridish containing three different concentrations (16.6, 33.3 and 83.3 mg/ ml) of hexane, chloroform, ethyl acetate, ethanol, and aqueous extract solutions. Each petridish was placed with 3 worms and observed for paralysis (or) death. The mean time for paralysis was noted when no movement of any sort could be observed, except when the worm was shaken vigorously; the time death of worm (min) was recorded after ascertaining that worms neither moved when shaken nor when given external stimuli. In the same manner albendazole was included as reference compound. The test results were compared with Reference compound Albendazole (15mg/ml) treated samples. The anthelmintic activity was performed according to the method (Ghosh et al., 2005).

RESULT AND DISCUSSION:

Phytochemical analysis was carried out on the plant Adhatoda vasica which revealed the presence of medicinally important bioactive compounds. The presence of phytochemical compounds in the plant Adhatoda vasica were evaluated in leaf, stem and root by using different solvents such as hexane, chloroform, ethyl acetate, ethanol and aqueous.

Phytochemical constituents of Adhatoda vasica:

Results obtained for the qualitative phytochemical screening of leaf, stem and root of Adhatoda vasica by using different solvents such as hexane, chloroform, ethyl acetate, ethanol and aqueous are presented in Table 1. Of the 10 phytochemicals screened, all the 10 compounds such as quinones, coumarins, terpenoids, phenols, saponins, flavonoids, volatile oils, resins, gums and mucilages and tannins were found present in the leaf of Adhatoda vasica than the other parts. Sagar Vijayrao Kathale (2013) studied the phytochemical screening of ethanolic leaf extract of Adhatoda vasica and reported that the leaves of Adhatoda vasica contain phytochemicals such as alkaloids, phenolics, tannins, flavanoids, saponins etc. Less separation of compound is recorded from stem and root extracts. The leaf extract of Adhatoda vasica contains more compounds, so the result indicates that Adhatoda vasica leaf hold promises as source of pharmaceutically important phytochemicals.

The presence of flavanoids could be extremely helpful as flavanoids possess antiallergic, anti-inflammatory, antiviral and antioxidant activities (Bbosa et al., 2010). Flavanoids in the body are known to reduce the risk of heart diseases (Urquiaga and Leighton, 2000). In terms of anti-cancer activity, they inhibit the initiation, promotion and progression of tumors (Urquiaga and Leighton, 2000; Okwu, 2004).

Tannins have been used to cure diarrhea (De Wet, 2010). Tannins are reported to have various physiological effects like anti-irritant, antisecretolytic, antiphlogistic, antimicrobial and antiparasitic effects.

The phenolic compounds are one of the largest ubiquitous groups of plant metabolites (Singh et al., 2007). The presence of phenols is considered to be potentially toxic to the growth and development of pathogens (Okwu and Okwu, 2004).

Saponins are used as dietary supplements and nutriceuticals (Yu-Fen, 2010). Saponins has the property of precipitating and coagulating red blood cells. Some of the characteristics of saponins include formation of foams in aqueous solutions, hemolytic activity and cholesterol binding properties and bitterness (Okwu, 2004).

Quinones are aromatic rings with two or more ketone substitutions. The natural quinone pigments range in colour from pale yellow to almost black and there are over 450 known structures (Harborne, 1973). The anti-hemorrhagic activity of quinones may be related to its ease of oxidation in body tissues (Harris, 1963).

Terpenoids exhibit various important pharmacological activities i.e., anti-inflammatory, anticancer, anti-malarial, inhibition of cholesterol synthesis, anti-viral and anti-bacterial activities (Mahato and Sen, 1997).

The results obtained in this study thus suggest that the identified phytochemical compounds may be the bioactive constituents and this plant (Adhatoda vasica) is proving to be an increasingly valuable reservoir of bioactive compounds of substantial medicinal merit.

Table 1: Phytochemical Constituents of Adhatodavasica

No.	Tests	Leaf	Stem	Root
1	Quinones	+	+	+
2	Coumarins	+	–	–
3	Resins	+	+	+
4	Terpenoids	+	+	+
5	Phenols	+	–	–
6	Saponins	+	+	+
7	Flavanoids (Ferric chloride test)	+	+	+
8	Gums and mucilages	+	+	+
9	Tannins	+	+	+
10	Volatile oils	+	+	–

+ indicates presence of the phytochemical

– indicates absence of the phytochemical

Anthelmintic activity of leaf extracts of Adhatoda vasica:

The crude extract samples, which were used to evaluate anthelmintic activity, showed variable times at different concentrations. The crude extracts of ethanol showed the significant anthelmintic effect causing death of the worm at all the concentrations but the time of death was different in each case. However, when observed the response of worms in case of paralysis, there was significant variation among the results produced by the extracts at different concentrations like 16.6, 33.3 and 83.3mg/ml. The ethanol extract showed more significant effect on paralyzing the worms, in terms of paralysis time, at every concentration compared to that of hexane, chloroform, ethyl acetate and aqueous extracts. Similar observations were made in the anthelmintic activity as well.

The effect of extracts on the paralysis (or) helminthiasis of the worm, according to the results (Table 2 and Plate 2) may be indicated as ethanol > aqueous > chloroform > ethyl acetate > hexane extracts. In particular the ethanol extract exhibited an increased paralytic as well as helminthiatic effect over albendazole at the given experimental concentrations (Table 2). This may be due to the increased level of extraction of tannins in ethanol followed by aqueous > chloroform > ethyl acetate > hexane extracts. The data presented in the table and observations made thereof, lead to the conclusion that the different degree of helminthiasis of the different extracts are due to the level of tannins present in compounds.

Tannins, the secondary metabolite, occur in several plants have been reported to show anthelmintic property by several investigators (Athnasiadou et al., 2001; Waller, 1997; Yesilada et al., 1993). Tannins, the polyphenolic compounds, are shown to interfere with energy generation in helminth parasites by uncoupling oxidative phosphorylation (Martin, 1997) or, binds to the glycoprotein on the cuticle of parasite (Thompson and Geary, 1995), and cause death. Coming to the chemistry of nematode surface, it is a collagen rich extracellular matrix (ECM) providing protective cuticle that forms exoskeleton, and is critical for viability. The mammalian skin also consists largely of collagen in the form of fibrous bundles. In leather making industry, vegetable tannins are commonly used in the tanning operation of leather processing that imparts stability to collagen of skin matrix through its reactivity and hence make the collagen molecule aggregate into fibres. This results in the loss of flexibility in the collagen matrix and gain of mechanical property with improved resistance to the thermal (or) microbial/enzymatic attack. Similar kind of reaction is expected to take place between the nematode cuticle (the earth worm) and the tannin of Adhatoda vasica, possibly by linking through hydrogen bonding, as proposed in this study. This form of reactivity brings toughness in the skin and hence the worms become immobile and non-functional leading to paralysis followed by death. Hence further investigation and proper isolation of the active principles might help in the findings of new lead compounds, which will be effective against various parasitic infections.

Table 2: Anthelmintic activity of leaf extracts of Adhatoda vasica

		No	Solvent	Concentration (mg/ml)	Time taken for paralysis (minutes)	Time taken for death (hours)
		1	Hexane	16.6	59	67.13
				33.3	57	67.04
83.3	55	66.54
		2	Chloroform	16.6	58	66.18
				33.3	55	66
				83.3	43	18.11
		3	Ethyl acetate	16.6	50	67
				33.3	48	66.45
				83.3	45	66.23
		4	Ethanol	16.6	46	18.26
				33.3	45	18.15
				83.3	42	18
		5	Aqueous	16.6	49	19
				33.3	46	18.48
				83.3	41	18.37
		6	Control	15	43 hours	57

Conclusion:

From the present study, it was concluded that among the various solvents used, the ethanolic leaf extract of Adhatoda vasica showed significant anthelmintic activity. This may be due to the presence of bioactive compounds. Thus the present study suggests that the extracted phytochemicals are very valuable. The wormicidal activity of various extracts of whole plant of Adhatoda vasica suggests that it is effective against parasitic infections of humans. Further, in future it is necessary to identify and isolate the possible active phytoconstituents responsible for the anthelmintic activity and study its pharmacological actions. Further extension of this research is possible in near future to find out the natural antibiotics from this plant origin.

REFERENCES:

1. Ahmed John, S. and M. Koperuncholan, 2012, Antibacterial activities of various solvent extracts from Impatiens balsamina, International Journal of Pharma and Biosciences., (3): 401-406.

2. Siddharthan, S., 2007, Systematic evaluation of natural phenolic antioxidants from 133 Indian Medicinal Plants, Food Chem., 102: 938-953.

3. Madhulika, A., 2010, In- vitro cytotoxicity of extracts and fraction of Calotropis procera (Ait) roots against human cancer cell lines, Int. J. Green Pharm., 4:36-40.

4. Raju, G., 2007, Indian medicinal plants as a source of antimicrobial agents, J Ethnopharmacol., 110 (2): 200-234.

5. Grover, J. K., 2002, Medicinal plants of India with antidiabetic potential, J. Ethnopharmacol., 81:81-100.

6. Marcy, J., 2005, Drug discovery from medicinal plants, Life Sci., 78:431-441.

7. Evans, M., 1994, A guide to herbal remedies, Orient Paperbacks.

8. Agarwal, A., 2005, Pharma Times., 37(6): 9-11.

9. Kokate, C. K., 1994, Practical Pharmacognosy, (Vallabh Prakashan, New Delhi), 1:15-30.

10. Criagg, G. M. and J. N. David, 2001, Natural product drug discovery in the next millennium, J. Pharm. Biol., 39: 8-17.

11. Tiwari. P., B. Kumar, M. Kaur, G. Kaur and H. Kaur, 2011, Phytochemical screening and extraction: A review, Internationale Pharmaceutica Sciencia., 1: 98-106.

12. Harborne, J. B., 1973, Phytochemicals Methods, Chapman and Hall Ltd., Londen, 8(9): 49-188.

13. Harborne, J. B., 1973, Phytochemical methods, A guide to Modern Techniques of Plants analysis, John Wiley and Son Inc., New York, 1-26.

14. Okwu, D. E., 2004, Phytochemicals and vitamin content of indigenous species of southeastern Nigeria, J. Sustain Agric Environ., 6(1): 30-37.

15. Edeoga, H.O., D.E. Okwa and B.O. Mbaebia, 2005, Phytochemical constituents of some Nigerian medicinal plants, African J. Biotechnol., (7): 685-688.

16. Lena, G., 2010, Phenolic compounds, antioxidant activity and in vitro inhibitory potential against key enzymes relevant for hyperglycemia and hypertension of commonly used medicinal plants, herbs and spices in Latin America, Bio Resour Technol., 10: 4676-4689.

17. Uniyal, S.K., K.N. Singh, P. Jamwal and B. Lal, 2006, Traditional Use of Medicinal Plants Among the Tribal Communities of Chhota Bhangal, Western Himalaya, J. Ethnobiol Ethnomed., 2:14.

18. Calixto, J.B., 2000, Efficacy, safety, quality control, marketing and regulatory guidelines for herbal medicines (phytotherapeutic agents), Braz. J. med. biol. Res., 33: 179-189.

19. Grover. J.K., S. Yadav and V. Vats, 2002, Medicinal Plants of India with Anti-diabetic Potential, Journal of Ethnopharmacology. 81(1): 81-100.

20. Mishra, L.C., B.B. Singh and S. Dagenais, 2000, Scientific Basis for the Therapeutic Use of Withania somnifera (Ashwagandha): A Review, Altern. Med. Rev., 5(4): 334-346.

21. Chaturvedi, M., S. Dwivedi, A. Dwivedi, P.K. Barpete and R. Sachan R, 2009, Formulation and Evaluation of Polyherbal Anthelmintic Preparation, Ethnobot., Leaflet, 13: 329-331.

22. Monteiro, A.M., S.W. Wanyangu, D.P. Kariuki, R. Brain, F. Jackson and Q.R. Mckellar, 1997, Pharmaceutical quality of anthelmintics sold in Kenya, Vet. Rec., 142: 396-398.

23. Waller, P.J., 1997, Anthelminthic resistance, Vet. Parasitol., 72: 391-405.

24. Waller, P.J., 1997, The global perspective of anthelmintic resistance in nematode parasites of sheep — excluding Australia, Proc. 4th Intl. Cong. Sheep Vet., Armidale, Australia, 59–63.

25. Kelly, J.D. and C.A. Hall, 1979, Resistance of animal helminths to anthelmintics, Adv. Pharm. Ther., 16: 89-128.

26. Okon, E.D., R.A. Ogunsusi and J.P. Fabiyi, 1980, Survey and feasibility studies on fascioliasis and parasitic gastroenteritis of ruminants in Nigeria, Federal Livestock Department of Nigeria Report, Lagos; 1-55.

27. Taylor, M.A. and K.R. Hunt, 1989, Anthelmintic drug resistance in the UK, Vet Rec., 125: 143-147.

28. Coles, G.C. and R.T. Roush, 1992, Slowing the spread of anthelmintic resistant nematodes of sheep and goats in the United Kingdom, Vet. Rec., 139: 505-510.

29. Geert, S. and P. Dorny, 1995, Anthelmintic resistance in helminthes of animals of man in the tropics, Bulletin-des-Seances, Academic-Royale-des-Sciencesd. Dutre Mer., 3:401-23.

30. Coles, G.C., 1997, Nematode control practices and anthelmintic resistance on British sheep farms, Vet.Rec., 141:91-93.

31. Satyavati, G.V., M.K. Raina and M. Sharma, 1976, Medicinal Plants of India, Vol. I, Indian Council of Med. Res., New Delhi, India, pp: 201–206.

32. Akhthar, M.S., Z. Iqbal, M.N. Khan and M. Lateef, 2000, Anthelmintic activity of medicinal plants with particular reference to their use in animals in the Indo-Pakistan subcontinent, Small Ruminant. Research., 38: 99-107.

33. Maciel, M.V., S.M. Morais, C.M.L. Bevilaqua, A.L.F. Camurca-Vasconcelos, C.T.C. Costa and C.M.S. Castro, 2006,Ovicidal and larvicidal activity of Melia azedarach extracts on Haemonchus contortus, Veterinary Parasitology., 140: 98-104.

34. Aslam, M., S. Rais, M. Alam and A. Pugazhendi, 2013, "Adsorption of Hg (II) from Aqueous Solution Using Adulsa (Justicia adhatoda) Leaves Powder: Kinetic and Equilibrium Studies", Journal of Chemistry., 1–11.

35. Patel, V.K. and H. Venkata-Krishna-Bhatt H, 1984, In vitro study of antimicrobial activity of Adhatoda vasica (L) (Leaf extract) on gingival inflammation, A preliminary report, Indian Journal of Medical Sciences., 38:70-72.

36. Trease, G. E. and M. D. Evans., 1989, A Textbook of Pharmacognosy Builler Tindall and Caussel London, 13^thedn. pp. 176-180.

37. Sofowara, A. 1993, Phytochemical screening of medicinal plants and traditional medicine in Africa, 2^nd Edition Spectrum Books Limited, Nigeria; 150-156.

38. Harborne, J. B., 2005, Phytochemical Methods, Springer (India) Pvt. Ltd., New Delhi, p. 17.

39. Kumar, A., R. Ilavarasan, T. Jayachandran, M. Decaraman, P. Aravindhan, N. Padmanaban and M. R. V. Krishnan., 2009, Phytochemical investigation on a tropical plant, Pak. J. Nutri., 8: 83-85.

40. Ghosh, T., T.K. Maity, A. Bose and G.K. Dash, 2005, Anthelmintic activity of Bacopa monierri, Indian, J.nat. Produc., 21;16-19.

41. Sagar, V.K., 2013, Phytochemical constituents in medicinal plants, Int. J. Pharma. Bio. Sci., 4(1): 930.

42. Bbosa, G.S., 2010, Medicinal Plants used by traditional Medicines Practitioners for the treatment of HIV/AIDS and related conditions in Uganda, Journal of Ethnopharmacol., 130:43-53.

43. Urquiaga, I. and F. Leighton, 2000, Plant polyphenol, antioxidants and Oxidative stress, Biol. Res., 33:159-165.

44. De Wet, H., 2010, Medicinal plants used for the treatment of diarrhoea in northern Maputaland, KwaZulu-Natal Province South Africa, J. Ethnopharmacol., 130: 284-289.

45. Singh, R., S. K. Singh, and S. Arora., 2007, Evaluation of antioxidant potential of ethyl acetate extract/fractions of Acacia auriculiformis, A. Cunn Food Chem. Toxicol., 45: 1216-1223.

46. Okwu, D. E and Okwu, M. E, 2004, Chemical composition of Spondias mombin Linn. plant parts, J. Sustain Agric Environ., 6(2): 140-147.

47. Yu-Fen, C., 2010, Foam properties and detergent abilities of the Saponins from Camellia oleifera, Int. J. Mo. Sci., 11: 4417-4425.

48. Harris, R. S., 1963, Vitamins K, p. 192-198, In M. Florkin, and E. Stotz (ed.), Pyrrole pigments, isoprenoid compounds and phenolic plant constituents, vol. 9. Elsevier, New York, N. Y.

49. Mahato, S.B. and S. Sen, 1997, Advances in triterpenoid research, 1990-1994, Phytochemistry., 44:1185–236.

50. Athnasiadau, S., I. Kyriazakis, F. Jackson and R.L. Coop, 2001, Direct Anthelmintic effects of condensed tannins towards different gastrointestinal nematodes of sheep: in vitro and in vivo studies, Vet. parasitol., 99: 205-219.

51. Yesilada, E., G. Honda, E. Sezik, M. Tabata, K. Goto and Y. Ikeshiro, 1993, Traditional medicine in Turkey IV, Folk medicine in the Mediterranean subdivision, Journal of Ethnopharmacology., 39: 31-38.

52. Martin, R.J., 1997, Mode of action of anthelmintic drugs, Vet. J., 154:11-34.

53. Thompson, D.P. and T.G. Geary, 1995, The structure and function of helminth surfaces in Biochemistry and Molecular Biology of parasites (j.j.marr, ed.), 1st ed. Academic Press, New York, 203-232.

Received on 09.11.2016 Modified on 15.12.2016

Research J. Pharm. and Tech. 2017; 10(2): 414-420.

DOI: 10.5958/0974-360X.2017.00083.X


Percolation- Hexane leaf extract	Percolation- Chloroform leaf extract		Percolation- Ethyl acetate leaf extract

Percolation- Ethanolic extract	Percolation- Aqueous extract		Control

Test- Hexane extract Test	Chloroform extract Test		Ethyl acetate extract

Test- Ethanolic extract		Test- Aqueous extract