PHYTOCHEMICAL AND ANTICONVULSANT STUDIES OF METHANOL LEAF EXTRACT OF HYMENOCARDIA ACIDA, TUL (EUPHORBIACEAE)

ABSTRACT

The research was on the phytochemical and anticonvulsant studies of methanol leafextract of Hymenocardia acida, Tul, (Euphorbiaceae) used inNorthern Nigeria for the treatment of headache, rheumatic pain, sickle cell crisis, malaria, epilepsy and cancer. The preliminary phytochemical screening of crude methanol extract (CME) using standard methods revealed the presence of terpenoids, tannins, saponins, alkaloids and flavonoids.Phytochemical evaluations were carried out using silica gel column chromatography, preparative thin layer chromatography and gel filtration using sephadex LH-20. The CME was partitioned successively with n-hexane, chloroform, ethyl acetate and n-butanol to yield different fractions. Extensive phytochemical investigation of n-Hexane soluble fraction of the leaf using silica gel column chromatography, gel filtration and preparative thin layer chromatography led to the isolation of Lupeol. The structure of the isolated compound was elucidated with the help of 1HMR and 13C NMR analysis. The oral median lethal dose (LD50)in mice was found to be greater than 5000mg/kg, suggesting the crude extract is practically non-toxic. Anticonvulsant activity was studied using maximum electroshock test (MEST) in chicks and pentylenetetrazole (PTZ) induced seizure model in mice. The CME of Hymenocardia acida at doses of 150mg/kg, 300mg/kg and 600mg/kg did not exhibit significant activity against MES convulsion because none of the chicks was protected against the seizure but there was 90% protection with the standard drugPhenytoin at a dose of 20mg/kg while the extract produced a dose independent activity in the PTZ induced seizure in mice which was significant at (p<0.05) which was seen as percentage protection against seizure as 50, 33.33, 16.67% at doses of 150mg/kg, 300mg/kg and 600mg/kg respectively i.e the lower the dose of the extract the higher the protection. The standard control, Sodium Valproate 200mg/kg protected the mice 100%.

The finding of the study suggests that the CME of Hymenocardia acida possesses significant anticonvulsant activity which might be due to the phytochemical constituents. This provides some scientific rationale for the ethnomedicinal claim of the use of the plant in the management of epilepsy.

CHAPTER ONE

1.0 INTRODUCTION

1.1 Natural Product

Natural Products can be defined as organic compounds and other chemicals synthesized by plants through metabolic processes aided by sunlight, involving CO2, H2O vapour and chlorophyll. Generally, natural products are characterized by specific functions they perform in plants and animals. Classical natural product chemistry methodologies enabled the discovery of a vast array of bioactive secondary metabolites from various source materials including terrestrial plants, terrestrial micro-organisms, marine organisms, and terrestrial vertebrates and invertebrates (Tyler et al., 1988). Natural products have been used since ancient times and in folklore for the treatment and prevention of many diseases and illnesses. They are the most successful source of potential drug leads (Haefner, 2003; Butler, 2004; Cragg andNewman, 2005; Mishra and Tiwari, 2011;Rey-ladino et al., 2011).

Therefore, natural product continue to provide unique structural diversity in comparison to standard combinatorial chemistry, and presents opportunities for discovering mainly novel low molecular weight lead compounds. Since less than 10% of the world’s biodiversity has been evaluated for potential biological activity, many more useful natural lead compounds await discovery with the challenge being how to access this natural chemical diversity (Cragg andNewman, 2005).

The earliest records of natural products were depicted on clay tablets in cuneiform from Mesopotamia (2600 B.C.) which documented oils from Cupressus sempervirens (Cypress) and Commiphora species (myrrh) which are still used today to treat coughs, colds and inflammation (Cragg andNewman, 2005). The Ebers Papyrus (2900 B.C.) is an Egyptian pharmaceutical record, which documents over 700 plant-based drugs ranging from gargles, pills, infusions to ointments (Cragg andNewman, 2005). The Chinese Materia Medica written sometimes around 1100 B.C. “Wu Shi Er Bing Fang” contains 52 prescriptions, Shennong Herbal (~100 B.C.) contain 365 drugs and the Tang Herbal (659 A.D) gave the records of 850 drug swhich are documented records of the uses of natural products (Cragg andNewman, 2005). The Greek physician, Dioscorides, recorded the collection, storage and the uses of medicinal herbs, whilst the Greek philosopher and natural scientist, Theophrastus (~300 B.C.) dealt with medicinal herbs (Cragg andNewman, 2005).

During the Dark and Middle Ages the monasteries in England, Ireland, France and Germany preserved this Western knowledge whilst the Arabs preserved the Greco-Roman knowledge and expanded the uses of their own resources, together with Chinese and Indian herbs unfamiliar to the Greco-Roman world (Cragg andNewman, 2005).

It was the Arabs who were the first to privately own pharmacies in the 8th century with Avicenna, a Persian pharmacist, physician, philosopher and poet, contributing much to the sciences of pharmacy and medicine through works such as the Canon Medicinae (Cragg andNewman, 2005).

1.2 Traditional Medicine

Traditional medicine refers to health practices, knowledge and beliefs incorporating plants, animals and mineral based medicines, spiritual therapies, manual techniques and exercises applied singularly or in combination to treat, diagnose and prevent illnesses or maintain well-being (WHO, 2005; NNMDA, 2008).The world Health organization estimated that 80 percent of people worldwide still rely on plant-based traditional medicines for some aspect of their primary health care (Farnsworth and Soejarto, 1985).

Medicinal plants are plants containing substances which can be used for medication or as precursor of drug synthesis (Sofowora, 1982). Medicinal plants can be referred to as: ‘all higher plants that have been alleged to have medicinal properties, i.e. effects that relate to health, or which have been proven to be useful as drugs by western standards, or which contain constituents that are used as drugs’ (Farnsworth and Soejarto, 1991). The term ‘medicinal’ as applied to a plant indicates that it contains a substance or substances which modulate beneficially the physiology of sick mammals, and that it has been used by man for that purpose (Fellows, 1991). Medicinal plants have been a source of medicine to human health since ancient time, whereas about 60-75% of world populations require plant for carrying health (Farnsworth, 1994; Joy et al., 1998; Harvey, 2000). Plants and microbes are the main source of natural products (Hayashi et al., 1997; Armaka et al., 1999; Lin et al.,1999a ;Lin et al.,1999b: Basso et al., 2005), and consistently become main source of the newest drugs (Harvey 2000). Many methods of investigation or drug development from natural sources are based on the bioassay-guided isolation of natural products on traditional uses of local plants (Ataur Rahman and Choudhary 1999). Ayurveda is the most ancient health caresystem and is practiced widely in India, Srilanka and other countries (Chopra and Doiphode, 2002). Atharvveda (around 1200 BC), Charak Samhita and Sushrut Samhita (100 – 500 BC) are the main classics that give detailed descriptions of over 700 herbs (Dash et al., 2001). In the western world, documentation of use of natural substances for medicinal purposes can be found as far back as 78 A.D., when Dioscorides wrote “De Materia Medica”, describing thousands of medicinal plants (Tyler et al., 1988).

This treatise included descriptions of many medicinal plants that remain important in modern medicine, not because they continue to be used as crude drug preparations, but because they serve as the source of important pure chemicals that have become mainstays of modern therapy.

The knowledge associated with traditional medicine (complementary or alternative herbal products) has promoted further investigations of medicinal plants as potential medicines and has led to the isolation of many natural products that have become well known pharmaceuticals. Many modern pharmaceuticals have been modelled on or derived from chemicals found in plants. An example is the heart medication digoxin (I) derived from foxglove (Digtialis purpurea), quinine (II), an antimalarial agent isolated from the bark of Cinchona succirubra tree. Paclitaxel (III) (Taxol®), a drug used for breast cancer was isolated from the bark of Taxus brevifolia (Pacific Yew) (Cragg, 1998). Taxol® is present in limited quantities from natural sources, its synthesis (though challenging and expensive) has been achieved (Cragg, 1998). It is widely known in ethnomedicine that various parts of a plant can possess different healing properties, for instance, the bark of Rauwolfiamombasiana is used for the treatment of malaria. The root of the same plant is used for the treatment of fever and anxiety states.The root, stem and leaves of another species of this plant, R. vomitoria, are used for fever (Iwu, 1993).

Morphine (IV) used for pain relief was derived from Papaver somniferum; a potent antimalarial drug named Artemisinin (V) was isolated from Artemisia annua as a remedy against the multidrug resistant strains of Plasmodium.