PHYTOCHEMICAL SCREENING AND ANTHELMINTIC ACTIVITY OF ROOT EXTRACT OF HUGONIA MYSTAX LINN AGAINST PHERETIMA POSTHUMA – AN IN VITRO STUDY

Inse rt graf ical abst ract figu re here Abstract. Back ground: Helminth infections are among the most common infections in man in developing countries they pose a large threat to public. These infections can affect most population in endemic areas with major economic and social consequences. Objective: In this paper, an ethnomedicinal plant, Hugonia mystax Linn. was evaluated for its preliminary phytochemical screening and in-vitro anthelmintic activity. Method: The earthworms resembled the intestinal roundworm parasites of human beings both anatomically and physiologically and hence where used to study the anthelmintic activity. The worms were acclimatized to the laboratory condition before experimentation. Results: Preliminary phytochemical screening showed the presence of various classes of secondary metabolites such as flavonoids, phenols, saponins, steroids, tannins and terpenoids. Anthelmintic activity of ethanolic extract of root showed significant activity. The anthelmintic activity revealed the medicinal potential of H. mystax to develop a drug against various human ailments. Conclusion: The present study brings detailed pharmacognostical profile of roots of Hugonia mystax L. The species, H. mystax has a restricted global distribution, occurring only in India and Sri Lanka. It is an unexplored medicinal plant in the Indian medicinal system. According to ethnobotanical information, the root powder of H. mystax is used as the best antidote, anthelmintic, febrifuge and for the treatment of peptic ulcers.


Introduction (optional)
Ayurveda, the ancient healing system of India, grow luxuriantly from the Vedic period in India. In history, the classical texts of Ayurveda like Charaka samhita and Sushruta samhita were written around 1000BC. Medicinal plants like turmeric, ashwagandha, ginger, brahmi, manjistha and tulsi are integral part of ayurvedic medicines. All these plants have been used for the treatment of diseases, traditionally and their secondary metabolite constituents are the sources of important modern drugs such as MOL2NET, 2018, 4, http://sciforum.net/conference/mol2net-04 2 atropine, codeine, digoxin, morphine, quinine and vincristine. According to one estimate nearly 70 % of the synthetic drugs have been derived from medicinal plants. Herbal medicine plays vital role in maintaining the health and wealth of mankind. Majority of world population use herbal medicines (1).
The World Health Organization (WHO) reports that approximately 21,000 plants have been used for medicinal purposes. Herbs have stood the test of time for their safety, efficacy, cultural acceptability and minimal side effects. Therapeutic power of some plants is mainly due to the presences of some secondary metabolites, which collectively are referred to as phytochemicals. These phytochemicals have potential to be developed as herbal medicines or could serve as precursors for modern medicine.
It is now widely understood that free radicals are involved in the pathogenesis of many diseases (2).
Helminthiasis is an infection disease caused by nematode worms such as Ascaris lumbricoides, Trichuris trichiura, Nectator americanus and Ancyclostoma duodenale. Infection occurred when ingesting food contaminated eggs or larvae, hands or utensils or through penetration of the skin by infective hookworm larvae in contaminated soil. Helminth infections are among the most common infections in man, affecting a large proportion of the world's population (3). In developing countries they pose a large threat to public health and contribute to the prevalence of malnutrition, anemia, eosinophilia, and pneumonia. Although the majority of infections due to worms are generally limited to tropical regions, they can occur to travellers who visited those areas and some of them can develop in temperature climate (4). Helminthiasis is a disease in which a part of the body is infested with worms such as pinworm, roundworm, or tapeworm. Typically the worms reside in the gastrointestinal tract but may also burrow into the liver and other organs; infected people excrete helminth eggs in their faeces, which then contaminate the soil in areas with inadequate sanitation. However, increasing problems of development of resistance in helminths against anthelmintics have led to the proposal of screening medicinal plants for their anthelmintic activity. The plants are known to provide a rich source of botanical anthelmintics (5). As we know very well, now a days the medicinal preparation available in the market from which most of them either not effective up to the mark or has to develop resistance resulting in reoccurrence again. Plant derived drug serve as a prototype to develop more effective and less toxic medicines. Helminth infection is a medical and public health problem of high magnitude, both in humans and domestic animals, causing considerable suffering and poor growth. In addition, helminths of livestock pose a serious economic loss, in particular in areas where extensive grazing is practiced (6,7).
Anthelmintics or anti helminthics are drugs that expel parasitic worms (helminths) from the body. They may also be called vermifuges (stunning) or vermicides (killing). An anthelmintic will destroy or expel worms from the digestive system. Helminthic infections are among the most common infection in human beings, affecting a large proportion of the world's populations. Diseases caused by helminth parasites in livestock continue to be a major productivity constraint, especially in small ruminants in the tropical and subtropical countries (8).

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The genus Hugonia L. of family Linaceae comprise about 40 species in the world; of which Hugonia mystax L. was reported from India. This plant Hugonia mystax is locally known as Modirakanni. Ethnobotanically, the fruits are used by the tribals of Kalakad Mundanthurai for the treatment of Rheumatism. Roots were used as anthelmintic, astringent and also used for dysentery, snake bite, fever, inflammation and rheumatism. Biological activities such as analgesic, antiinflammatory and ulcerogenic were also reported. Roots of Hugonia mystax were evaluated for preliminary phytochemical screening and anthelmintic activity. Preliminary phytochemical screening showed the presence of various classes of secondary metabolites such as flavonoids, phenols, saponins, steroids, tannins and terpenoids.
Taking into consideration of medicinal value and utility, the present study was planned to explore anthelmintic potential of the medicinal plant named H. mystax (9,10).
The aim of present study is to investigate the anthelmintic activity of ethanolic extract of root of Hugonia mystax Linn against Pheretima posthuma (in vitro study).

Materials and Methods (optional) Plant material
Fresh roots of Hugonia mystax was collected from surroundings of Medchal dist., Telangana, India.

Preparation of extract
The root of Hugonia mystax was collected, washed with running tap water, shade-dried at room temperature and grounded in a manual mill to get a coarse powder of 60 mesh. Powdered plant materials of Hugonia mystax was extracted with 80% ethanol in a soxhlet apparatus at 40 0 C.
Extraction was done with solvent until the supernatant in the soxhlet apparatus became transparent (for 48 hours). The extracts were filtered through a Buchner funnel with whatman filter paper no. 1. The filtrate was evaporated to dryness under reduced pressure using rotary evaporator at 40 0 C. The crude MOL2NET, 2018, 4, http://sciforum.net/conference/mol2net-04 5 extract was stored at 4 0 C in airtight bottles in refrigerator. The ethanolic extract of the root was used for anthelmintic activity.

Qualitative phytochemical evaluation
The different chemical tests were performed for establishing profile of the extract for its chemical composition; the following chemical tests for various phytoconstituents in the ethanol extract was carried out as described below. Appearance of pink colour indicates the presence of terpenoids.
ii) Hirshonn reaction: When the substance was heated with trichloroacetic acid, red to purple colour was observed.   Paralysis was said to occur when the worms do not revive even in normal saline. Death was concluded when the worms lost their motility followed with fading away of their body colour.
Observations were made for the time taken to paralysis and death of individual worms.

Results and Discussion (optional)
Preliminary Phytochemical analysis of ethanolic extract of root of Hugonia mystax was presented in Table 1. Phytochemical studies have revealed the presence of several phytochemicals including alkaloids, glycosides, flavonoids, steroids, proteins, phenolic compounds, anthraquinones and tannins.   The preliminary phytochemical analysis of the extract has shown the presence of phenolics, like tannins and flavonoids as well as anthraquinones. The function of the anthelmintic drugs, like piperazine citrate, is known to cause paralysis of the worms so that they are expelled in the feaces of man and animals. Piperazine citrate by increasing chloride ion conductance of worm muscle membrane produces hyper polarization and reduced excitability that leads to muscle relaxation and flaccid paralysis. The extracts not only demonstrated this property, but they also caused death of the worms. Synthetic phenolic anthelmintics like niclosamide, interfere with the energy generation in the helminth parasites by uncoupling the oxidative phosphorylation. Another possible mechanism of action is that they bind to free proteins in the gastrointestinal tract of the host animal or to glycoprotein on the cuticle of the parasite and by this cause death. Tannins have also been shown to produce anthelmintic activities. There are reports for anthelmintic property of phenolics present in plant extract like Hugonia mystax root.

Conclusions (optional)
The traditional claim of root of Hugonia mystax as an anthelmintic has been confirmed as the extract shown activity against Pheretima posthuma. Further studies are necessary to isolate and reveal the active compound contained in the crude extract of Hugonia mystax responsible for activity and to establish the mechanism of action. Based on the present study results it can be used for the development of new pharmaceutical drugs for treatment and curing of helminthiasis and also this study shows that these extract offer a safe method or supplement treatment strategy to control helminthiasis.
However, further detailed study is needed to isolate and purification of constituents from the plant for anthelmintic activity.