Pharmacological and Therapeutic Activities of Kigelia africana (Lam.) Benth

Occurring widely in Africa and beyond is Kigelia africana (Lam.) Benth, a medicinal plant with several attributes and considerable potentials. Various parts of the plant are used locally to treat cancer, ulcer, gynecological disorders, genital infections, skin diseases, diabetes, epilepsy, bacterial and fungal infections as well as being used as cosmetics. The antioxidant and antiinflammatory properties of some parts of the plant have been explored for therapeutic purposes. Phytochemical analyses revealed the presence of a wide range of secondary metabolites. Toxicological effects of different extracts of the fruit, stem bark and leaf have been reported. In this review we provide an up-to-date information on established pharmacological and toxicological properties, as well as phytochemical constituents responsible for these activities in Kigelia africana.

The tree can grow up to 20m tall. It is evergreen where rainfall occurs throughout the year, but deciduous where there is a long dry season. The flowers (and later the fruit) hang down from branches on long flexible stems. Flowers are produced in panicles; they are bell-shaped, orange to reddish or purplish green and about 10cm wide. Their scent is most notable at night indicating their reliance on pollination by bats, which visit them for pollen and nectar [4]. Kigelia africana is characterized by its huge fruits which can weigh between 5 to 10kg [5]. The fruit is indehiscent, with woody wall and heavily marked with lenticels at the surface. It is grey-brown and many seeded when matured. The fruit is eaten by several species of wild animals that also disperse the seeds in their dung [4]. Parts of the plant are used for treating a wide range of ailments traditionally based mainly on cultural practices. The fruit is used to treat skin ailments like fungal infections, boils, psoriasis and eczema. Dysentery, ringworm, tapeworm, postpartum haemorrhage, malaria, diabetes and pneumonia are also treated with the fruit [3]. The fruit is also applied for the treatment of solar keratosis and malignant melanoma [6]. The bark is used to treat venereal diseases while the root is applied to treat ulcer [3].
This review on Kigelia Africana is divided into biological activities such as antiprotozoal, antibacterial, antifungal activities; pharmacological properties such as antiinflammatory and analgesic, anticancer, antidiarrhoeal, anti-ulcer effects as well as toxicological effects with the responsible phytochemicals.

Anti-Protozoal Activity
One of the several uses of Kigelia africana is for treating malaria [7]. In-vitro studies revealed the efficacy of hexane, dichloromethane, ethyl acetate and ethanol extracts of the root bark against Plasmodium falciparum [8] and Trypanosoma brucei brucei and T. b. rhodesiense [9], the causative organisms for malaria and sleeping sickness respectively. The growth of Entamoeba histolytica was also inhibited by the stem bark butanol extract [10]. Four compounds that exhibited significant antiplasmodial activity were isolated from the ethyl acetate extract of Kigelia africana. Three of the four compounds showed good activity against all the different parasite strains, the chloroquineresistant W-2 and two field isolates of Plasmodium falciparum, with IC 50 <5µM. Specicoside exhibited the highest activity on W-2 (IC 50 =1.5µM) followed by 2β, 3β, 19α-trihydroxyurs-12-en-28-oic acid (IC 50 =1.60µM) and atranorin (IC 50 =4.41µM) while p-hydroxycinnamic acid was the least active (IC 50 =53.84µM) [11]. Lapachol in the methanol extract of the root and another compound (a quinone) obtained from the wood show anti-malarial activity. Three iridoidsspecioside, verminiside and minecoside isolated from the butanol extract of the stem bark possess ant-amoebic activity [7]. The antitrypanosome activity of the stem bark and root bark extracts are attributed to 2-(1-hydroxyethyl)naphtho-[2,3-b]-furan-4,9-quinone and three naphthoquinoids: isopinnatal, kigelinol and isokigelinol [9].

Antibacterial and Antifungal Activities
Various parts of Kigelia africana are employed to treat bacterial and fungal infections. In a study to verify these properties, crude extracts of stem bark and fruits were prepared with distilled water, ethanol or ethyl acetate. In the microtitre plate bioassay, the stem bark and fruit extracts showed similar antibacterial effects against Gram-negative and Gram-positive bacteria [12]. A mixture of three fatty acids exhibiting antibacterial effects was isolated from the ethyl acetate extract of the fruits using bioassayguided fractionation. Palmitic acid was the major antibacterial compound in this mixture thus supporting the traditional use of the plant in therapy of bacterial infections [12]. A biologically monitored fractionation of the methanolic extracts of the root and fruits led to the isolation of the naphthoquinones, kigelinone, iso-pinnatal, dehydro-α-lapachone and lapachol and the phenylpropanoids, ρ-coumaric acid and ferulic acid as the compounds contributing to the observed antibacterial and antifungal activities [11].
In another antibacterial and antifungal study using the agar diffusion technique, Owolabi and co-workers [13] reported that like amoxillin standard antibiotics, crude ethanolic extract exhibited antibacterial and antifungal activities against Staphylococcus aureus and Candida albicans with zones of inhibition measuring 15.0±0.95 and 20.75±4.6 mm respectively but the aqueous extract exhibited no antibacterial or antifungal activity. The minimum inhibitory concentration for the ethanol extract was found to be 6.25±1.07mg/ml for S. aureus and 7.92±1.52 mg/ml for C. albicans. In another report, it was also established that the stem bark extract inhibited a number of harmful microorganisms, including Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus and Candida albicans [14]. Similarly, evaluation of the antibacterial activities of ethanolic and aqueous extracts of Kigelia africana fruit against multi drug resistant Pseudomonas aeruginosa showed that the ethanolic extract was more potent than the aqueous extract [15].

Anti-Inflammatory and Analgesic Activities
The use of the bark, stem, twigs, leaves and fruits of Kigelia africana to relieve rheumatism, toothache and headache has been documented [16]. Picerno and co-workers [17] reported that the anti-inflammatory property of Kigelia africana fruit polar extract was due to the constituent verminoside. The compound is known to cause significant anti-inflammatory effects inhibiting both iNOS expression and NO release in the LPS-induced J774.A1 macrophage cell line.
The ethanolic extract of the stem bark has been evaluated for analgesic property using acetic acid induced mouse writhing and hot plate reaction time; and anti-inflammatory property using the carrageenan-induced paw oedema. The extract showed a dose dependent significant reduction of the number of writhes with 500mg/kg body weight dose giving the highest reduction. In the carrageenan-induced paw oedema, a dose dependent significant inhibition was observed (p<0.001) between the second and the fifth hour, confirming that the ethanolic stem bark extract has significant analgesic and anti-inflammatory properties. Inhibition of the synthesis of prostaglandins and other inflammatory mediators has been suggested to be responsible for the analgesic and anti-inflammatory properties [18].

Anti-Diarrhoeal Activity
One important local use of Kigelia africana is the use of the leaf for treating diarrhea [19]. An administration of a dose of 100 or 200 mg/kg of aqueous leaf extract to experimental animals caused anti-diarrhea activity. It also reduced the reduced fecal output in castor oil -induced diarrhea in animals and remarkably decreased the propulsive movement of the gastro-intestinal contents [2]. On the isolated guinea pig ileum, the extract did not appreciably affect acetylcholine and histamine induced contractions. In an antidiarrhoeal activity studied using castor oil to induce diarrhea in rats (in vivo) and using isolated jejunum, 500 and 1000mg/kg ethanol root extract (in vitro) significantly reduced the frequency of diarrheal stool and the spontaneous propulsive movement of isolated jejunum [2]. Kigelia africana root extract also produced reversible inhibition of acetylcholine induced mobility of isolated rabbit jejunum. The observed spasmolytic effects of the extract may explain its continual use in the management of chronic abdominal pains associated with diarrhea [2].

Anti-Diabetic and Antioxidant Activities
Like many other African food [20] and medicinal plants [21][22][23][24], the use of Kigelia africana to manage diabetes is traditionally practiced and reported [2,3]. The anti-diabetic activity and the antioxidant effect were studied [25]. Also, in a polyherbal preparation, ADD-199, Kigelia africana is in combination with three other plants: Maytenus senegalenses, Annona senegalensis and Lannea welwitchii. The anti-diabetic and antioxidant effects were investigated in streptozotocin-induced diabetic C3H mice and results were compared with two allopathic hypoglycaemic drugs, glibenclamide and metformin. Plasma glucose, insulin and lipids as well as liver glycogen and lipid peroxidation were measured following treatment for eight weeks. The results indicated that plasma insulin levels in normal controls at termination were about 76µmol/L compared to trace levels in untreated diabetic mice. Like glibenclamide, ADD-199 increased insulin levels in diabetic mice up to 70% of levels in untreated non-diabetic mice whilst metformin had no effect. Also, basal plasma glucose levels in diabetic controls (18.8mM) were reduced to 14.0mM by 100mg/kg ADD-199 in < 2 weeks compared to 4 to 6 weeks for glinbenclamide and metformin, respectively. This hypoglycaemic effect of ADD-199 was associated with the alkaloidal content of the extract. Treatment with ADD-199 or the hypoglycaemic agents reversed the observed elevation in plasma lipids but increased hepatic glycogen, triacylglycerol and cholesterol levels. Treatment also increased hepatic glucose uptake by isolated diaphragms and attenuated hepatic lipid peroxidation. These antihyperglycaemic and antioxidant actions of ADD-199 were comparable to those of the maximum daily therapeutic doses of glibenclamide (0.25mg/kg) and metformin at 50mg/kg [25].
Olaleye and Rocha, [26] carried out an ex-vivo assessment of the antioxidant property of Kigelia africana extracts in rat liver homogenate. Administration of different pro-oxidants: 10 µM iron (II) sulphate, (FeSO 4 ) , 5µM sodium nitroprusside (SNP), and 2mM 3-nitropropionic acid led to increased formation of thiobarbituric acid reactive substances (TBARS), which indicates lipid peroxidation in the liver. Administration of Kigelia africana statistically (p<0.05) reduced the production of TBARS in a concentration-dependent manner in all the prooxidant-induced oxidative stress, suggesting that the use of the plant in the treatment of various diseases, especially liver diseases could be due to its ability to act as an antioxidant [26]. Saini and co-workers [3] attributed the antioxidant potential of Kigelia africana to caffeic acid derivatives and other compounds unique to the plant.

Anti-Ulcer Effect of Kigelia africana
The use of Kigelia africana fruit, bark and root to treat ulcer has been reported [3]. Owolabi and Nworgu [27] investigated the anti-ulcer activity of the ethanol extract of Kigelia africana stem bark in Wistar albino rats. In both preventive and curative models of ulcer respectively induced by absolute ethanol and indometacin, the extract caused marked inhibition of ulceration, suggesting a dose-dependent gastro-protective effect by the plant in the two models of ulcer [27].

Acute toxicity
In a study on the diuretic activity of aqueous extract of the bark in experimental rats, Sharma and colleagues [28] reported that it was safe up to 5g/kg. A determination of acute toxicity of the methanol fruit extract using male Sprague-Dawley rats showed that the extract was well tolerated by the animals as there were no observable signs of acute toxicity effects like restiveness, seizure or dizziness after the administration of 400mg/kg. However at 6400mg/kg, the animals showed signs of toxicity like jerks and writhes with 60% death. At 12,800mg/kg there was 80% death of the animals. The LD 50 was estimated from a logdose curve to be 3,981.07 mg/kg [29][30][31].
In another study, 100mg/Kg aqueous extract was administered to rats induced with acetaminophen liver toxicity. The extract countered the effect of acetaminophen on the activities of aspartate transaminase (AST), alanine transaminase (ALT), superoxide dismutase (SOD), catalase (CAT), gluthathione peroxidase (GPx) and δaminolevulinate dehydrogenase (δ-ALA-D). This suggests that the extract can act as hapatoprotective agent against toxicity possibly through its antioxidant action [26].

Sub-chronic and chronic toxicity
The administration of the aqueous anti-diabetic polyherbal extract ADD-199 containing Kigelia africana and three other plants at a daily dose of 100 or 500mg/kg body weight over 30 days, to male Wistar albino rats appeared to show no effect on many haematological, urinary and plasma biochemical parameters. It also had no effect on some modulators of some hepatic cytochrome P450 (CYP) isozymes normally measured as indices of organ specific toxicity or potential for drug interactions. Specifically ADD-199 containing Kigelia africana did not affect plasma AST, ALT, alkaline phosphatase (ALP) and albumin or creatinine kinase (CK) levels [25]. It also did not affect plasma creatinine and urea levels. Furthermore, ADD-199 neither affected packed cell volume (PCV), nor the levels of red blood cells (RBC), reticulocytes, platelets, lymphocytes and granulocyte. It however, caused significant dose-dependent reductions in white blood cell counts at day 15 with varying degrees of recovery by day 30. ADD-199 also reduced the rate of body weight increases after week 3. However, no changes were observed in organ weight at termination. The ADD-199 did not significantly affect zoxazolamine-induced paralysis and pentobarbital-induced sleeping times as well as certain CYP isozyme activities in rats, suggesting that ADD-199 had no overt organ specific toxicity and did not demonstrate a potential for drug interactions via CYP-mediated metabolism in rats following sub-chronic administration [25].
The protective effect of methanol extract of Kigelia africana fruit extract against cisplatininduced renal toxicity in male rats has been studied [29]. The rats treated with cisplatin for 28 days, suffered loss in body weight, elevation in blood urea nitrogen and serum creatinine levels as well as tubular necrosis. Pre-treatment with Kigelia africana fruit methanol extract as a prophylaxis significantly prevented these changes. Though post-treatment of animals with the extract after cisplatin treatment did not completely restore serum catalase activity, it caused some alleviating effects, suggesting that Kigelia africana fruit extract may protect against cisplatin-induced renal toxicity, and hence might serve as a novel agent to limit renal injury [29].

Cytotoxic activity
The cytotoxicity of hexane, chloroform, ethyl acetate, ethanol and methanol extracts of different parts of Kigelia africana has been studied on Artemia salina using the brine shrimp lethality test (BSLT). Some workers [31] have reported moderate toxicity of the ethanol extract of the root and fruit at a dosage of 593 and 124µg/ml respectively while the ethyl acetate extract of the fruit was also moderately toxic at 495µg/ml. Other workers [31,32] reported a moderate cytotoxicity of ethanol extract of the fruit to Artemia salina at a dosage of 1000µg/ml.

Molluscidal and Piscicidal Effects
The molluscidal and piscidal effects of the aqueous extract of Kigelia africana bark has been reported [33].

Anticancer Activity
There are many reports in literature suggesting the use of Kigelia africana to either prevent or to treat cancer [4,34,35]. In a study to determine the effect of Kigelia africana seed oil on cell proliferation in culture, human colon adenocarcinoma (Caco-2) and human embryonic kidney (HEK-293) cells were maintained and treated with various concentrations (0, 20, 40, 80, 100 and 120mg/l) of Kigelia africana seed oil. The trypan blue dye exclusion method was used to determine cell growth 48 hours after oil treatment. The seed oil suppressed both Caco-2 and HEK-293 cell growth in a dose dependent manner. The seed oil did not cause increase cell death as the number dead cells remained unchanged under control and oil-treated conditions. The oil significantly suppressed Caco-2 cell growth compared to HEK-293 cell growth at all oil concentrations. The suppression of Caco-2 and HEK-293 cell proliferation by Kigelia africana seed oil suggest a potential antiproliferative effect of the oil on the two cell lines [36].
Methanolic extract of the root of Kigelia africana contains the constituent lapachol [34] which is reported to be effective in the treatment of solar keratosis, skin cancer and Kaposi sarcoma, an HIV-related skin ailment [35]. Serial dilutions of standardized aqueous, ethanol and dichloromethane extracts of the stem bark and fruits of Kigelia africana were tested for their growth inhibitory effects against four melanoma cell lines and a renal cell carcinoma line (Caki-2) using two different assays (3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide, MTT and Sulforhodamine B, SRB assays). Lapachol, a possible constituent of these extracts, together with known therapeutic anti-neoplastic agents evaluated this way, showed significant inhibitory activity of the dichloromethane extract of the stem bark and lapachol in a dose-dependent and timedependent manner. Chemosensitivity of the melanoma cell lines to the stem bark was greater than that seen for the renal adenocarcinoma line, but in marked contrast sensitivity to lapachol was similar amongst the five cell lines, suggesting that lapachol is the active ingredient that exhibit anti-cancer property [37].

Effect on the Central Nervous System
Among the variety of uses of Kigelia africana is in the treatment of epilepsy, CNS stimulating activity and as antidotes against snake poisons. Snake bite antidotes are made with an infusion of the fruits, stem, leaves, twig or bark taken orally or rubbed onto the bite [38]. The CNS stimulant activity of the ethanolic stem bark extract has been verified [39]. The barbiturate induced sleeping time and the Rota rod bar were used to study the effect of the extract on muscle coordination in mice. The results showed that the extract at all doses tested reduced the duration of sleeping time when compared to the control group that received distilled water. This difference in sleeping time was significant (p<0.0001 at all doses tested) and was found to be dose dependent. Its effect was also compared with caffeine (a known stimulant) and the extract gave a shorter duration of sleeping time compared to caffeine (p<0.05 at 400 mg/kg dose) indicating better stimulant properties. In comparison with diazepam, the extract at all doses tested also gave a statistically significant shorter duration of sleep (p<0.0001). On the Rota rod, the extract had no sedative effect as the animals maintained their balance on the rod through the entire period of the experiment [39].

Effects on Animal Reproductive Organs and Reproductive System
In traditional medicine, sexual complaints such as infertility, poor libido, sexual asthenia and impotence are treated with herbal prescriptions containing the fruit, roots or leaves of Kigelia africana. A small amount of unripe fruit is chewed or an aqueous preparation of the fruit is taken orally as a sexual stimulant, and the intoxicating traditional beer to which they are added is drunk as an aphrodisiac [5]. The fruits are also applied on the breast to improve flow of milk in lactating women [3]. Kigelia africana fruit aqueous extract has been successfully used as fertility enhancing agent in rats [40]. The steroidal components are thought to enhance reproductive ability since steroids as androgen and estrogen have shown to contain fertility properties necessary for the improvement and production of reproductive organs [41]. A study to investigate the effects of varying dietary supplementation of Kigelia africana on the sperm quality and fertility in African catfish, Clarias gariepinus showed that dietary inclusion of the plant positively affected some parameters of sperm quality in the fish, with increases in sperm counts, percentage motility, milt volume and motility duration [41].

Use as Cosmetics
Traditionally, Kigelia africana is used as cosmetic to enhance beauty [5]. Some preparations contain extract of one or more parts of the plant, mainly the fruit, stem bark or the pendulum (where the fruit hangs from, or a product thereof). Typically, the preparation contains 50% extract mixed with carrier, excipients and colorants (Sumobrain TM ). Aqueous or alcohol extracts are ideal for water based cosmetic products such as gels, lotion, water or oil emulsions and creams. The products are used to make anti-ageing and regenerating skin care products, skin tightening cosmetics such as bust firming products. Antiinflammatory, antioxidant and antibacterial agents are other products that are commercially made from Kigelia Africa.

Diuretic Activity
The diuretic activity of Kigelia africana aqueous bark extract was investigated by the determination of urine volume, electrolyte concentration and diuretic potency in male albino rats. Different concentrations of the extract, 250 and 500mg/kg were orally administered to hydrated rats and their urine output was immediately measured after 5 hours of treatment. Fusemide (10mg/kg) was used as reference drug while normal saline (0.9%) was used as control. The result showed that the bark extract exhibited dose dependent diuretic property. The onset of diuretic action was within 1 hour and lasted up to 5 hours, with 500mg/kg displaying a potency of 0.8 and 250mg/kg, respectively. The extract also caused a marked increase in Na + , K + and Cl labels [28]. The result suggests that the aqueous extract possess significant diuretic activity, justifying its use in folk medicine for kidney and urinary disorders.

NUTRITIONAL VALUE
Kigelia africana provides a nutritious source of food during times of famine when the seeds are roasted to eat. The fruit and bark are used in the brewing process to aid fermentation and enhance the flavor of traditional beer. The fruit pulp is not edible as it may cause blistering of the tongue and skin. However, fallen fruits along with leaves and flowers are browsed or foraged by livestock and wildlife [5]. In a study of the effect of Kigelia africana fruit meal (KAFM) on sperm quality of African catfish, Clarias gariepinus, the KAFM supplement may have enhanced nutrient utilization which is reflected by improvement in weight gain by testes [40].
The pro-fertility effect of dried fruit meal (KAFM) was investigated on reproductive performance of female Clarias gariepinus fed with increasing levels for 90 days in relation to egg production and quality (number, shape, structure, fecundity) and hatchability (percentile fertilization, percentile hatching, percentile survival

PHYTOCHEMISTRY OF VARIOUS PARTS Kigelia africana
The occurrence of secondary metabolites in different parts of Kigelia africana is responsible for its several medicinal applications. These compounds include naphthaquinones, iridoids, sterols, coumarins, flavonoids and alkaloids among others [3,43,44] Structures of isolated compounds have been characterized and identified using gas chromatography, ultraviolet spectroscopy, infrared spectroscopy, nuclear magnetic resonance and other spectroscopic techniques like, mass spectrometry, or gas chromatography-mass spectrometry.
The use of Kigelia africana in traditional African medicines has been verified by corresponding pharmacological properties.

Chemical Constituents of Kigelia Africana Stem
A study of the antimicrobial properties of the aqueous stem bark extract of Kigelia africana revealed the presence of two naphthoquinones kigelinone and isopinnatal [44,3].Three known iridoids: specioside, verminoside and minecoside have also been isolated from the stem bark [47] The dichloromethane extract of the stem bark contain naphthoquinones which possess anti trypanosomal properties [3,4] while kigelin, βsitosterol, 1,3-dimethylkigelin and ferulic acid have been isolated from the bark, while the isolation of kigeliol from the wood and balaphonin from the stem bark have been reported [19,48].

Chemical Constituents of Kigelia Africana Leaf
The hexane extract of the leaf of Kigelia africana has been reported to be rich in hydrocarbons and some volatile compounds. In a study that qualitatively and quantitatively analyzed the hexane extract for various chemical compositions, it was revealed to contain twelve compounds with the major ones identified as nhentriacontane, 1-tricosene, 11-(2,2dimethylpropylheneicosane, 2,6,10trimethyldodecane, pentafluoroheptadecyl ester, 2-ethylhexyloctadecyl sulfurous acid ester, heneicosane and hexyloctylsulfurous acid ester [55,56,57] Others are 4,4-dimethylundecane, methyl-12-methyltetradecanoate, 1iodohexadecane and 1-iododecane. Hentriacontane have been reported to have a possible anti-tumour activity while methyl-12methyltetradecanoate has also been reported for its inhibition capacity on the development of coneal angiogenesis, which is responsible for blindness and other infections [44] Flavonoids and iridoids [45] and a 7-O-glucoside [9,37] have also been found in the leaves ( Table 2). The structural formulae of some of the chemical constituents isolated from Kigelia africana are presented in Fig. 1.
Considering the pharmacological and therapeutic efficacies of many phytochemicals isolated from different parts of Kigelia africana in experimental models, it would be worthwhile to synthesize the active components in large quantities for testing in higher animals like monkeys and chimpanzees preparatory to large scale clinical trials in humans before their deployment as therapeutic agents in humans. However, since the synthesis of many phytochemicals can be unbelievably difficult [21], it is recommended that this plant be included as protected species in communities where they are found, and plantations of this plant be established as immediate source of the identified active ingredients for the clinical trials.