Antifungal Susceptibility Pattern of Seed Exracts of Ricinodendron Heudelotii (njangsa) on Candida Albicans and Aspergillus Niger

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DOI : https://doi.org/10.52793/ACMR.2024.5(3)-82

Table1: Some fungi of medical importance.

Diseases caused by Candida albicans and Aspergillus niger

Candidiasis

Candida albicans is the commonest cause of candidiasis (moliniasis). The yeast is a common commensal of gastrointestinal tract. Most Candida infections are opportunistic, occurring in debilitated persons. Candidiasis is also associated with prolong broad spectrum antibiotic therapy [20]. Many different clinical forms of candidiasis are known, involving primarily the mucosal surfaces (thrush), gastrointestinal or urogenital tracts and deep-seated infections such as candideamia or meningitis. Candida vaginitis is a common infection during pregnancy [20]. Candida infections of the mouth and oesophagus occur in 80-95% of those with HIV disease [20].

Aspergillosis

Aspergillus niger is a saprophyte mould. Most live in the environment without causing disease. Disease caused by aspergillus include allergic bronchopulmonary aspergillosis (causes asthma and eosinophilia) due to hypersensitivity to aspergillus antigen inhaled in airbone conidia; aspergilloma in which inhaled conidia germinate in a pulmonary cavity and grow into a “fungus ball” and; invasive aspergillosis in which the fungus infects the lung (causing acute pneumonia) and spread to other organs, producing abscesses and necrotic lesions. Invasive aspergillosis occurs mainly in immunocompromised patients and is often life threatening [20].

Ricinodendron heudelotii

Geographical distribution of Rrcinodendronheudelotii

Ricinodendron heudelotii is common in tropical regions. The native geographic location of “njangsa” reaches from Senegal in West Africa to Sudan, Uganda and Tanzania and from Sudan down to the western coast of Sub-Sahara Africa to Angola. The tree is also found in Madagascar and Cameroon where the main production area is the humid forest zone [21].

Ricinodendron heudelotii grows generally in rain forests and is also typical for secondary forests. This tree is a light- demanding species [21]. Therefore, it can also be found in deciduous forests, forest edges, secondary scrubs and in semi-dry savannahs. The tree is observed in food crop fields, cocoa farms and other agro-forestry systems, where the tree can also intentionally be planted [9].

Taxonomy

Kingdom: Plantae; Phylum: Tracheophyta; Class: Magnoliopsida; Order: Malpighiales; Family: Euphorbiaceae; Sub-Family: Crotonoideae; Tribes: Ricinodendreae; Genus: Ricinodendron; Species: R. heudelotiis; Binomial name: Ricinodendron heudelotii; Subspecies: R heudelotii heudelotii R heudelotii africanum; [22].

Synonyms

Jatropha heudelotii baill [22].

Common names

French: boisjasanga, essang, essessang.

English: African nut-tree, African oil-nut-tree, African wood oil-tree, ezang, njangsa/ njangsang, Ricinodendron [22].

Vernacular names

In Angola, it is called Mugella; In Cameroon, it is called Essessang; In Zaire it is called Bofeko; In Ghana, it is called Wama; In Nigeria, it is called Okhuen; In Uganda, it is called Kihongo; In Coted’ivoire, it is called Akpi [21].

Description of Ricinodendron heudelotii

Ricinodendron heudelotii is a dioecic plant found in central and West Africa. The tree is fast growing and reaches a height between 20 and 50m with straight trunk which can have a diameter up to 2.7m. His crown is broad and the roots are widely spread. The bark is smooth with a grey color. Inside, the bark is red when cut [21].

The flowers are yellowish-white, 5mm long and form a long terminal panicle which measures between 15 and 40cm. Flowering time is between April and May. Male panicles are longer and slender than female flowers [21].

Njangsa trees produce fruits that are typically 2-3 lobed and contain 2 cells in which the seeds lie. These seeds are red-brown to black in color, rounded and some 1cm in diameter. The seeds are oily in texture and can be bought either raw or dried. They have an odor reminiscent of oily chocolate, but their flavor is truly unique: subtly aromatic with a mild bitter taste [21]. At maturity (August- September) the fruits smell like over-ripe apples [21].

How to Extract the njangsa Seeds?

Extracting the njangsa seeds from the fruits can be quite demanding. The big green kidney-shaped fruits are normally gathered into piles under the tree. The fruits are left for up to four weeks to decompose so that the fleshy parts can easily peel off [23].

The nuts are then extracted from the rotten fruits after which they are boiled for a long time. Once the nuts are a bit softened, they can then be cracked for the inner kernels to be dried. The dried njangsa kernels can be kept for years and used as desired [23].

Chemical Constituents of Ricinodendron heudelotii seeds

The medicinal plants are useful for healing as well as for curing human diseases because of the presence of phytochemical constituents. Phytochemicals are non- nutritive chemicals that have protective or disease preventive properties [24]. They are non-essentials nutrients present in plants leaves, roots, fruits, vegetables and back of trees. A compositional analysis of “njangsa” revealed a presence of long chain omega-3 fatty acids not usually associated with plant material. The seed has 31.4% crude protein and 44.7% lipid [25]. Of this lipid, about73% is composed of polyunsaturated fatty acids, almost entirely of eicosapenteanoic acid, with about 18% oleic acid [25].

The seed also contains about 8.9% of crude fibers, 5.6% of carbohydrates with 495kcal being the energetic value per 100g of seeds [9]. The phytochemical analysis of the oil extracts from the seeds showed the presence of the following minerals: Na, K, Ca, Mg, Mn, Fe, Cu and Zn as well as the following family of compounds: steroids, saponins, and terpenoids (Mordi set al, 2016). Phytochemicals play many rules in the body.

Functions of phytochemicals in the body

Researchers have found that phytochemicals have the potential to stimulate the immune system, prevent toxic substances in the diet from becoming carcinogenic, reduce inflammation, prevent DNA damage and aid DNA repair, reduce oxidative damage to cells, slow the growth rate of cancer cells, trigger damage cells to self- destruction (apoptosis) before they can reproduce, help regulate intracellular signaling of hormones and gene expressions, and activate insulin receptor [26,27]. Some major functions are:

1. Antioxidant: Most phytochemicals have antioxidant activity and protect our cells against oxidative damage and reduce the risk of developing certain types of cancer. Phytochemicals with antioxidant activity: allyl sulfides (onions, leeks, garlic), carotenoids (fruits, carrots), flavonoids (fruits, vegetables), polyphenols (tea, grapes) (Biesalki, 2008).
2. Hormonal action: Isoflavones, found in soy, imitate human estrogens and help to reduce menopausal symptoms and osteoporosis (Biesalki, 2008).
3. Stimulation of enzymes: Indoles, which are found in cabbages, stimulate enzymes that make the estrogen less effective and could reduce the risk of breast cancer. Other phytochemicals, which interfere with enzymes, are protease inhibitors (soy and beans), terpenes (citrus fruits and cherries) (Biesalki, 2008).
4. Interference with DNA replication: Saponins found in beans interfere with the replication of cell DNA, thereby preventing the multiplication of cancer cells. Capsaicin, found in hot peppers, protects DNA from carcinogens (Biesalki, 2008).
5. Anti-bacterial effect: The phytochemical allicin from garlic has anti-bacterial properties (Biesalki, 2008).
6. Physical action: Some phytochemicals bind physically to cell walls thereby preventing the adhesion of pathogens to human cell walls. Proanthocyanidins are responsible for the anti-adhesion properties of cranberry. Consumption of cranberries will reduce the risk of urinary tract infections and will improve dental health (Biesalki, 2008).

Brief study of some secondary metabolites

Alkaloids

Alkaloids are complex compounds, basic in nature, defined by the amine function, which provides its constituent chemical properties that are related to a high toxicity. Alkaloids belonging to beta-carboline group possess antimicrobial, anti- HIV and anti-parasitic activities [28]. They also act as proteins synthesis, protective substances against animals and insects, detoxicating agents of the plant [29].

Flavonoids

Flavonoids are the pigments that color most flowers, fruits, and seeds. They are secondary metabolites, widely distributed in plants [30]. Flavonoids are powerful antioxidants with anti-inflammatory and immune system benefits.

Glycosides

Glycosides are basically molecules where in a sugar is attached to a non- carbohydrate element, generally a minor natural molecule. They are soluble in water and alcohol and have several important functions in all living organisms. A number of plants stock up compounds in the form of dormant glycosides. These chemicals may be set in motion by certain enzymes by means of hydrolysis that results in the sugar portion to be detached. The activation of these chemicals by the enzymes makes them accessible for utilization. A number of such glycosides enclosed by the plants are of therapeutic value [31].

Tannins

Tannins are water-soluble polyphenols that are commonly found in higher herbaceous and woody plants [32] mainly in leaf, bud, seed, root, and stem tissues. They can be classified into two categories: hydrolysable and non- hydrolysable (condensed). Tannic acid is an important gallotannin belonging to the hydrolysable class, while catechin belongs to the non-hydrolysable class. Hydrolysable tannins are esters of phenolic acids and a polyol, usually glucose [33]. It has been reported that tannic acid and propyl gallate, but not gallic acid, were inhibitory to foodborne bacteria, aquatic bacteria, and off- flavor-producing microorganisms. Their antimicrobial properties seemed to be associated with the hydrolysis of ester linkage between gallic acid and polyols hydrolyzed after ripening of many edible fruits. Tannins in these fruits thus serve as a natural defense mechanism against microbial infections. They inhibit the growth of many fungi, yeasts, bacteria, and viruses [33]. They also play a role in protection from predation, and perhaps also as pesticides, and in plant growth regulation [34].

Resins

Resins are "solid or highly viscous substances," which are typically convertible into polymers. Such viscous substances can be plant-derived or synthetic in origin. They are often mixtures of compounds. The medical action of resins vary enormously. Generally speaking, most resins are antimicrobial and wound healing in animals and in the plants that secrete them. Topical application of pine resins to wounds and burns has been shown to stimulate local immune function, normalized wound hemodynamics, and stimulate squamous epithelization. Beyond this, resins from different plants can have a wide array of effects. for example, frankincense, the gum resin of Boswellia serrata, is well documented as an inflammation modulator that is helpful in asthma and ulcerative colitis; this is attributed at least in part to its inhibition of 5-lipoxygenase. The gum resin of commiphora molmol (myrrh) is antiparasitic, analgesic, and antineoplastic.

Saponin

Saponins are glucosides with foaming characteristics which consist of polycyclic aglycones attached to one or more sugar side chains. The aglycone part, also called sapogenin, is either steroid (C27) or a triterpene (C30) [35]. The foaming ability of saponins is caused by the combination of a hydrophobic (fat-soluble) sapogenin and a hydrophilic (water-soluble) sugar part. Saponins have a bitter taste and are phytochemicals which can be found in most vegetables, beans and herbs. The best-known sources of saponins are peas, soybeans, and some herbs with names indicating foaming properties. Saponins have many health benefits. Studies have illustrated the beneficial effects on blood cholesterol levels, cancer, bone health and stimulation of the immune system.

Traditional uses of the tree

Roots, leaves, back and seeds of the tree have many medicinal uses and they are used in the treatment of ailments with the trunk (back) being the most effective and frequently used part. Many of the medicinal uses range from use as laxative, treatmentof diarrhea, cough, dysentery, aneamia, blennorrhoea, sexual and infertility problem [36].

Roots

The roots or root- back of the “njangsa” tree can be mixed with salt and bush pepper, which is used as a laxative. Laxative helps to stimulate the evacuation of fecal wastes from the bowels. Also, the roots can be mixed together with the back and the decoction used for the treatment of venereal diseases [8].

Back

The back is used by the pregnant women to relieve labour pain and the decoction can be taken by the new mothers to relieve stomachache after birth. It is also used by infusion to prevent miscarriage. The back decoction is used for the treatment of sexual problems, menstrual pains, blennorrhoea and infertility problems [8]. It is used for washing, treating and healing of wounds/sores as well as treatment of rheumatism, aneamia. The bark extract can serve as an antidote for neutralizing poisons: this is because the extracts contain lupeol. The back is also used for the treatment of coughs, catarrh, cold, diarrheoa and dysentery. “njangsa” bark can be ground, warmed and then used for treating elephantiasis [36].

Leaves

Leaves are used for the destruction of worms e.g: extracting guinea- worm. It is also used with the latex as a strong purgative [8].

Seeds

The seeds combined with palm oil to form lotion, can be rubbed on body’s soft spots. “njangsa” seeds can be used for preparing soups to stimulate appetite [36].

Other uses

“Njangsa” tree can be used as a natural manure to improve the soil because the roots are populated by mycorrhizae [9]. The bark, leaves, stems, woods and fruits of the “njangsa” tree can serve as a natural fertilizer. It can be cut and used as stakes for supporting other plants. Fallen njangsa fruits are attractive to animals thus hunters use the tree to guide animals to their game [8].

For constructive purposes, the light white wood of Ricinodendron heudelotii can be used for building and construction. The wood can be used for making toys, cutleries etc. the wood can also be used for electrical insulation. The wood can be carved into plates, spoons, bowls, ladles, pestles, mortars, platters and stools. It is also used for firewood purposes [8].

“Njangsa” seeds can be used as xylophones for generating musical sounds It is also used by villagers for playing local games [9]. The husks and seeds can be processed into oil. Oil processed from dried kernel and wood ash from the burnt “njangsa” tree is suitable for making soap, varnish and indigo dye [9]. The wood ash of the “njangsa” tree can be used as a cooking salt [8].

Materials

The materials used are:

• Fruit material: “njangsa” seeds.
• Instruments: Weighing boat, glass funnel, cotton wool, sieve, grinding machine, sieve, test tubes, Petri dishes, wire loop, measuring cylinder, micropipette, bijou bottles, universal bottles, syringes.
• Chemicals: 95% alcohol (ethanol), distilled water (500ml), Gram stain reagents, 1% aqueous HCl, 6% ferric chloride, iodine solution, concentrated HCl, ammonia solution, Benedict qualitative reagent, dilute NaOH, concentrated H2SO4.
• Test organisms: Candida albicans, Aspergilus. niger.,
• Culture media: Sabaroud Dextrose Agar.
• Equipment: Incubator, balance, microscope, autoclave, fridge, Bunsen burner.

Method

Description of the study area

This research was done in the Science for Life Foundation Laboratory created in 2006 as not-profit organization and located N05°57.921’, E010°09.370’ Nitob IV quarter’s Bamenda II. The mission is to faster community wellness, good environmental stewardship, alleviation of poverty and the building of sustainable live hoods among rural and urban poor. The laboratory bears number of units that is the bacteriology/parasitology, serology, biochemistry, hematology and physiotherapy.

Study design

An experimental research design was used to study the effects of njangsa extracts on C albicans and Aspergillus niger.

Study period

This research was carried out from the 18 january to the 27 Febuary 2017.

Sample preparation and extraction

Well dried njangsa seeds were collected during the dry season in food market Bamenda, it was then grind into powder form by means of a clean and dried machine.

Aqueous extraction

75g of the powder seeds was put in a separate clean and sterile 1000ml conical flask containing 250ml of distilled water. The flask was shaken intermittently for 24 hours. The material was then filtered using Watmann No1 filter paper and the filtrate was transferred into a clean, sterile evaporating dish and was evaporated to dryness in an oven at 500C.

Alcoholic extraction

75g of the powder seeds was put in a separate clean and sterile 1000ml conical flask containing 250 ml of ethanol. The flask was shaken intermittently for 24 hours. The material was then filtered using Watmann No1 filter paper and the filtrate was transferred into a clean, sterile evaporating dish and was evaporated to dryness in an oven at 500C.

Fractionation of the extract

2g of each extract was dissolved into 10ml of distilled water. This gives a concentration of 200mg/ml

Test organisms

Pure cultures of the test organisms were obtained from the Regional Hospital Bamenda (C albicans) and Science for Life Foundation (A niger) where they were identified on the bases of microscopical staining reaction and biochemical tests.