Ethnomedicinal uses, phytochemistry and pharmacological properties of the genus, Kirkia

Purpose: To review the phytochemical, ethnopharmacology and traditional uses of the Kirkia species throughout their distributional range. Methods: The information documented in this article is from scientific journals, books, theses and reports obtained from library collections and electronic search engines such as Google, Google scholar, publishing sites such as Elsevier, ScienceDirect, BioMed Central (BMC), PubMed and other scientific database sites such as ChemSpider and PubChem. Results: Kirkia species are used as herbal remedies for abdominal pain, cholera, cough, snake bites and toothache in East, Central and Southern Africa. There are similarities in the use of Kirkia species as herbal medicines in Central and Southern Africa in terms of plant parts used, herbal preparation, route of administration and dosage. The chemical composition of Kirkia species is dominated by fatty acids, flavonols, isocoumarin, lignans, neolignans, nor-carotenoids, phenols and tannins isolated from leaves, stem bark and roots. Major biological activities demonstrated by Kirkia species include antimicrobial, antioxidant, antiplasmodial and antiplatelet activities. Conclusion: Kirkia species are valuable herbal medicines with multiple pharmacological effects. Kirkia extracts and their isolates are potential sources of modern medicines following future detailed studies to elucidate their mechanisms of action, clinical trials.


INTRODUCTION
The monotypic genus Kirkia Oliver is a member of the dicotyledonous family Kirkiaceae Takhtajan which contains six species [1,2]. The genus Kirkia was named after Dr John Kirk (1832 -1922), an English doctor and plant collector, who accompanied Livingstone on his Zambezi expedition [3]. Genus Kirkia is distributed in tropical Africa from Ethiopia and Somalia, south to Madagascar, South Africa and Namibia [4,5]. Kirkia acuminata Oliver is the most known and widely distributed species in the genus [4,6]. It is semi-deciduous, monoecious medium-sized tree, drought resistant and generally grows in hot and dry areas of Angola, Botswana, Democratic Republic of Congo (DRC), Malawi, Mozambique, Namibia, Zambia, Zimbabwe and South Africa [7].
Kirkia burgeri Stannard is a shrub or small tree divided into two sub-species, K. burgeri subsp. burgeri growing mainly on limestone slopes in Ethiopia. Kirkia burgeri subsp. burgeri is categorized as vulnerable (VUB2ab) (i,ii,iii,iv) on the IUCN Red List [8], primarily because of reduction in population size based on decline in area of occupancy, extent of occurrence and/or quality of habitat. Kirkia burgeri subsp. somalensis Stannard is confined to gypsum and anhydrite slopes in northern Somalia where it is categorized as near threatened [9]. Kirkia burgeri subsp. somalensis is generally rare and threatened by habitat degradation, grazing and over-exploitation of the species as a source of construction timber. Kirkia dewinteri Merxm. & Heineis a small tree up to 10 m tall endemic to the limestone and dolomite outcrops in the northwestern part of Namibia [4,6,10].
Kirkia dewinteri is categorized as vulnerable (VUD2) on the IUCN Red List in Namibia [11] primarily because of habitat destruction. Ecological research done on K. dewinteri showed that the species is generally rare in Namibia, characterized by restricted distributional range and in need of protection from human disturbances and wild fires [10]. Kirkia leandrii (Capuron) Stannard, is a tree, endemic to dry deciduous forest of Bemaraha, central Madagascar [5]. Kirkia tenuifolia is a shrub or small tree which is common or occasional in sandy or stony bushland in Ethiopia, Kenya and Somalia [4]. Kirkia wilmsii Engl. is a deciduous tree endemic to South Africa, found on granite and dolomitic soils in dry bushveld areas or on rocky slopes in Gauteng, Limpopo, Mpumalanga and the North West provinces [4,6,12].
Preliminary chemical and pharmacological studies done so far on Kirkia species have shown promising results with the possibility that these phytochemical compounds may be responsible for the medicinal uses and biological activities demonstrated by the species. The genus certainly has a high use value. The present review compiles fragmented information on the phytochemical, ethnopharmacological and traditional uses of the Kirkia species, hoping that this information will highlight the ethnopharmacological importance of the genus and will provide baseline information for future researchers intending to do further work on the genus.

Vernacular names and traditional uses
Kirkia species are known by various vernacular names in different geographical areas in Eastern, Central and Southern Africa (Table 1). Insight into the societal value of Kirkia species in East, Central and Southern Africa can be gained by examining these vernacular names. A survey of literature showed at least 39 vernacular names for K. acuminata in Southern Africa (Table 1). Kirkia burgeri subsp. burgeri is known by two vernacular names in Ethiopia (Table 1), while Kirkia tenuifolia is known by one vernacular name in Ethiopia (Table 1). South Africa [6,17] Kirkia dewinteri is known by one vernacular name in Namibia (  Table 2). For example, bark infusion of K. acuminata is taken as a remedy for abdominal pains in South Africa [27] and Zimbabwe [21]. Root decoction is taken as remedy for cough in Botswana [13,14], Mozambique [28], South Africa [27] and Zimbabwe [21]. Fruit juice of K. acuminata is used as a snake bite antidote in Botswana [14] and Zimbabwe [21,23,24]. Such similarities may be ascribed to efficacy of the plant species. This observation is supported by Hossan et al [33] who argued that a plant species is regarded as effective herbal medicine if it is used by many people in different geographical regions for the same disease or ailment.
The most frequently used plant parts are bark (53 %), roots (23 %), fruits and leaves (12 % each) ( Figure 1A). It is well recognized by conservationists that medicinal plants primarily valued for their root parts and those which are intensively harvested for their bark often tend to be the most threatened by over-exploitation [30]. There is need for implementing conservation strategies and mechanisms in East, Central and Southern Africa aimed at conserving Kirkia species, given the fact that K. burgeri subsp. burgeri, K. burgeri subsp. somalensis and K. dewinteri are threatened with extinction [8,9,11].
The population decline of Kirkia species is due to habitat degradation and loss [8,9,11], naturally rare and highly localized in distribution [10,11], over-exploitation of the species [9] and medicinal plant trade. According to Moeng and Potgieter [31], K. wilmsii roots were observed in 44 % of the medicinal plant (muthi) markets in the Limpopo province, South Africa. Given the widespread usage of Kirkia species in traditional medicine resulting in negative impacts on wild populations calls for conservation strategies aimed at halting or reducing these negative impacts. Therefore, conservation of Kirkia species and the traditional knowledge associated with their medicinal applications are important for sustainable utilization of the species. It is important to take concrete conservation measures aimed at protecting the species against over-exploitation and habitat loss. Kirkia remedies are often utilized in the form of decoction or infusion (60.9 %), bark powder mixed with food (17.4 %), ointments or paste (13.0 %), body and tooth wash (4.3 %) ( Figure  1B). Most (70.6 %) of the preparations are prescribed orally ( Table 2). All Kirkia remedies are used as monotherapies ( Table 2).
In addition to the medicinal uses, the swollen roots of K. acuminata, K. tenuifolia and K. wilmsii are used as sources of water during drought [5,7,17,20,34]. Leaves, seeds and twigs of K. acuminata and leaves of K. burgeri subsp. burgeri and K. wilmsii are browsed by game and livestock [17,25,45]. The timber or wood of K. acuminata and K. wilmsii are used for construction poles, carving household utensils and furniture [12,40]. Fibre obtained from the bark, young shoots and roots of K. acuminata and K. wilmsii used for weaving [6,7,12,17]. Kirkia acuminata is grown around homestead boundaries as a barrier or hedge or live fence and also as an ornamental and shade tree. In South Africa, Zambia and Zimbabwe, K. acuminata is commonly used as live fence around fields, gardens, homesteads and schools as a form of protection against animals; around enclosures (kraals) for livestock. The poles of K. acuminata take root and grow easily [12] explaining the high proportion of study households (23 %) with the species growing around their homesteads as live fence in Malawi [41].
Fresh leaves and twigs of K. acuminata have a wide variety of nutrients such as minerals, fibre, proteins and fat [43,46,47]. Kirkia acuminata leaves, seeds and twigs are a good source of minerals such as calcium, copper, iron, magnesium, manganese, phosphorus, sodium and zinc [43,45,47]. According to Aganga and Mosase [43], K. acuminata seeds have adequate quantities of phosphorus, calcium, magnesium, potassium, iron and copper required for beef, sheep and goat growth and production, while the content of sodium, manganese and zinc are below t h e recommended levels required by ruminants for growth and productivity. Some of these nutrients are present in low concentrations, but may have significant impact on animal health as both K. acuminata and K. wilmsii are important stock feed in southern Africa [17,20,43,46,47].
Nooteboom [48] isolated quercetin, caffeic acid, ellagic acid and large quantities of gallic acid from hydrolysed leaf extracts of K. wilmsii ( Figure  2). Similarly, Gemeda and Hassen [50] isolated acetic acid, propionic acid, iso butyric acid, butyric acid and valeric acid from both K. Although there is a possibility that these phytochemical compounds may be responsible for the medicinal uses and biological activities demonstrated by both K. acuminata and K. wilmsii, but no studies were found in literature with details of biological activities of these compounds isolated from Kirkia species.

Pharmacological activities
Some of the pharmacological activities of K. acuminata and K. wilmsii reported in literature correlate with some of their ethnomedicinal uses listed in Table 2. While some of these pharmacological activities may not relate directly to their ethnomedicinal uses, they may provide some insight into potential therapeutic value and bioactive properties of K. acuminata and K. wilmsii. The biological activities that have been reported so far include antimicrobial [52][53][54][55], antioxidant [56], antiplasmodial [36] and antiplatelet [56] activities.  Timber, wood Wood used for making household utensils South Africa [12,40] Weaving Fibre obtained from the bark, young shoots and roots is strong and used for weaving South Africa [5,6,12,17]

Antimicrobial activity
One of the most common ethnomedicinal uses of Kirkia species, K. acuminata, K. tenuifolia and K. wilmsii is in the treatment of a wide range of infectious diseases caused by microorganisms. Such diseases or ailments include symptoms such as cholera [22,24,34], diarrhoea [22,24], dysentery [22,24], fever and malaria [33,36,39], toothache [4,7,20] and wounds [21,23,24]. Such wide use of Kirkia species against microbial infections in traditional medicine prompted Masoko [54] to evaluate antibacterial effects of acetone, dichloromethane, hexane and methanol leaf extracts of K. acuminata against Enterococcus faecalis, Escherichia coli, Pseudomonas aeruginosa and Staphylococcus aureus with ampicillin as the positive control. In this study, E. faecalis was the most sensitive microorganism with average MIC value of 0.1 mg/ml followed by E. coli (0.31 mg/mL) while S. aureus and P. aeruginosa were resistant with MIC values of 0.97 mg/mL) and 1.25 mg/ml respectively [54]. Similarly, Mmushi et al [52] evaluated acetone, dichloromethane, hexane and methanol leaf extracts of K. acuminata against Mycobacterium smegmatis using rifampicin as a positive control. The acetone, dichloromethane, hexane and methanol extracts demonstrated antimycobacterial activity with MIC values ranging between 0.31 to 0.63 mg/mL against M. smegmatis [52].
Suleiman et al [53,57] evaluated the hexane, acetone, dichloromethane and methanol extracts of K. wilmsii against Enterococcus faecalis, Escherichia coli, Pseudomonas aeruginosa and Staphylococcus aureus using the bioautographic procedure and a two-fold serial microdilution method. The authors found 18 inhibition bands against bacteria. Suleiman et al [53,57] also evaluated the hexane, acetone, dichloromethane and methanol extracts of K. wilmsii against Aspergillus fumigatus, Candida albicans, Cryptococcus neoformans, Microsporum canis and Sporothrix schenckii using the bioautographic procedure and a two-fold serial microdilution method. The authors found 11 inhibition bands against fungi. The acetone, dichlomethane, hexane and methanol extracts of K. wilmsii had antibacterial and antifungal activities with MIC values ranging from 0.07 to 2.50 mg/mL against the tested bacteria and fungi [53,57]. The leaves of the K. wilmsii further showed biological activity against the animal fungal pathogen Aspergillus fumigatus and the MIC ranged from 0.17 to 2.11 mg/mL [57]. In another study, Chigayo et al [55] evaluated the antimicrobial activity of aqueous root extracts of K. wilmsii using an HPLC gradient elution method with kanamycin as the positive control. The extracts showed antimicrobial activities against Aeromonas hydrophilia, Candida albicans, Enterobacter aerogenes, Escherichia coli, Proteus mirabilis, Salmonella typhi, Shigella dysenteriae, Staphylococcus aureus, Vibrio cholerae with minimum inhibitory concentration (MIC) ranging from 0.08 mg/mL to 3.445 mg/ml. Results of these studies implies that K. acuminata and K. wilmsii may be used as a broad spectrum antibiotics.

Antiplasmodial activity
Clarkson et al [36] evaluated leaf extracts of K. wilmsii against Plasmodium falciparum using the parasite lactate dehydrogenase (pLDH) assay. The dichloromethane/methanol (1:1) was highly active and showed promising antiplasmodial activity with IC 50 of 3.7 µg/mL, while water extra showed a weak activity with IC 50 of >100 µg/mL [36]. These pharmacological evaluations are important as the bark and leaf decoctions of K. wilmsii are used as herbal medicine for fever and malaria in South Africa [36,39]. Such pharmacological evaluations are also important for future research focusing on control and management of parasites and mosquito vectors in the tropics.

Antioxidant and antiplatelet activities
Extracts of K. wilmsii exhibited antioxidant properties in the qualitative assay using DPPH with EC 50 value of 3.57 ± 0.41 μg/mL [56]. In the quantification of antioxidation using ABTS, the extracts of K. wilmsii showed low antioxidant activities with respect to TEAC values of 0.67. Kirkia wilmsii showed low antiplatelet activity with EC 50 value of 0.22 ± 0.02 μg/mL [56]. The documented antioxidant and antiplatelet activities of K. wilmsii may be attributed to the presence of phytochemicals since compounds such as flavonols and phenolics are known to have antioxidant properties [58] and also lower the risk of cancer, heart diseases, hypertension and stroke [59]. Detailed anti-hypertensive studies of K. wilmsii are recommended based as ethnopharmacological studies by Semenya and Potgieter [37] revealed that 97 % of traditional healers in the Limpopo province, South Africa use K. wilmsii in a similar manner as hypertension remedy in terms of herbal preparation, route of administration and dosage strength.

Toxicity studies
Kirkia wilmsii extracts showed comparably low toxicity when compared with the reference agent berberine (cytotoxic agent) with hemagluttination assay titre value of 0.80 and agglutination value of 1.25 mg/mL [56]. Toxicological evaluations carried out so far on Kirkia are preliminary considering their widespread usage as herbal medicines in southern Africa. It is important to ascertain any toxicological effects that can occur as a result of chronic of sub-chronic usage of Kirkia species, and the different plant parts should be tested against a wide range of cell lines as well as using other in vitro toxicological assays and in vivo studies.

CONCLUSION
Of the six Kirkia species, only K. acuminata, K. wilmsii and to some extent K. tenuifolia have been studied so far under different ethnobotanical and ethnopharmacological aspects. The others are either threatened with extinction or are highly localized, naturally rare and in need of human intervention to save them from extinction. There is therefore need to develop conservation strategies and mechanisms for Kirkia species that are threatened with extinction as well develop micropropagation protocols as an alternative and viable means to provide sufficient plants to meet the herbal medicine needs and at the same time protecting the natural populations.
From literature, it is clear that both K. acuminata and K. wilmsii have been used for centuries in southern Africa as herbal medicines for microbial infections in humans and scientific studies carried out so far have validated some of their traditional antimicrobial uses. Up to now, only a few pharmacological studies directly dealing with the documented traditional uses of K. acuminata and K. wilmsii have been performed. Phytochemical studies led to the identification of a large number of fatty acids, flavonols, isocoumarin, lignans, neolignans, norcarotenoids, phenols and tannins. Most of these phytochemicals have not been evaluated, therefore, these compounds must be evaluated biologically in more detail. Further investigations should focus on phytochemical studies of these isolated compounds and try to link such pharmacological properties to documented ethnomedicinal uses.
Concerning toxicological information, no significant data are reported. A weak cytotoxic activity is described only for K. wilmsii [56]. Future research should focus on assessing biological and toxicological aspects of the leaves, stem bark and roots of Kirkia species used as herbal medicines as well as both their bioactive extracts and isolated compounds. There is no doubt that results presented in this review may provide useful clues to promote further investigations of ethnopharmacological properties as well as potential use of phytochemicals isolated from Kirkia species.

Acknowledgement
Financial support of this work by the National Research Foundation (NRF) and Govan Mbeki Research and Development Centre (GMRDC), University of Fort Hare is gratefully acknowledged.