Medicinal uses, phytochemistry, pharmacology and toxicological properties of Elaeodendron croceum

Elaeodendron croceum is a shrub or a small tree used as an emetic, purgative and herbal medicine for cough, sore throat, fever, tuberculosis, blood in sputum, chest congestion, human immunodeficiency virus (HIV) opportunistic infections and to clean the digestive tract. Phytochemical compounds identified from the species include alkaloids, cardiac glucoside, flavonoids, phenols, proanthocyanidins, saponins, sugars, tannins and triterpenoids. Pharmacological studies revealed that E. croceum extracts and compounds have antimycobacterial, anti-arthritic, antibacterial, anti-HIV, antioxidant, anti-inflammatory, antifungal and cytotoxicity activities. Future studies should focus on ethnopharmacological safety and efficacy of E. croceum extracts and compounds isolated from the species using in vivo preclinical studies and clinical trials.


INTRODUCTION
Elaeodendron croceum (Thunb.) DC. is a shrub or a small evergreen tree species which belongs to the Celastraceae family. The species is commonly referred to as forest saffron, saffron or wood saffron. E. croceum has in the past classified under several scientific names such as Cassine crocea (Thunb.) Kuntze, C. papillosa (Hochst.) Kuntze, Crocoxylon croceum Thunb. N. Robson, C. excelsum Eckl. & Zeyh., E. capense Eckl. & Zeyh., Salacia zeyheri Planch. ex Harv., Ilex crocea Thunb., E. croceum Thunb. DC. and E. papillosum Hochst. [1,2]. E. croceum is among the top 15 medicinal plants that are becoming scarce in KwaZulu Natal province in South Africa based on the information provided by traditional healers [3][4][5]. The could be as a result of overcollection as the species is widely sold in informal herbal medicine markets in both rural and urban areas of Gauteng [6] and KwaZulu Natal provinces in South Africa [4]. Overcollection of wild populations of E. croceum as a source for medicinal plant trade and other anthropogenic activities such as habitat destruction and land clearing for agricultural purposes have caused a severe reduction in the population size of the species [7]. E. croceum is regarded as Least Concern on the IUCN Red List of Threatened Species [8] but due to local conservation concern in South Africa as the population of the species is declining, the species has been flagged as declining following recommendations made by Williams et al [9], Von Staden et al [10] and Victor and Keith [11]. It is against this background that the medicinal uses, phytochemistry, pharmacology and toxicological properties of E. croceum were reviewed.

METHODS
Data on medicinal uses, botany, phytochemistry, pharmacological and toxicological properties of E. croceum were collected from multiple internet sources including Science Direct, SciFinder, Pubmed, BMC, Elsevier, Google Scholar, Scopus and Web of Science. Complementary information was gathered from pre-electronic sources such as books, book chapters, theses, scientific reports and journal articles obtained from the University library. The search for this information was carried out between May 2017 to August 2018. The keywords used in the search included "ethnobotany", "ethnomedicinal uses", "medicinal uses", "phytochemistry", "biological activities", "pharmacological properties", "toxicological properties", "Elaeodendron croceum", the synonym of the species and the English common names "forest saffron", "saffron" or "wood saffron". The internet search generated 562 articles in total. After duplicate articles and those with limited raw data were excluded, 36 articles were included in this study. These articles included 21 journal articles, books (nine), scientific report (three), websites (two) and a single book chapter.

FINDINGS AND DISCUSSION
Botanical profile and medicinal uses of E. croceum E. croceum ( Figure 1) grows up to a height of 15 metres with a trunk diameter of 250 mm [12]. The species has been recorded in montane forest margins and along the escarpment, mainly on seepage zones. The main stem is red-brown to grey in colour and may have some dark lenticels when young characterized by pale grey and thin bark, peeling off to reveal bright orange and yellow under-bark [13]. The leaves are opposite to sub-opposite in arrangement and oblong to elliptic in shape. The apex and base of the leaves is broadly tapering, leaf margins are strongly serrated and sometimes spined. The leaves are glossy green above and little paler below with prominent venation on both sides. The flowers are small, whitish or pale green in colour, occurring axillary clusters. The fruits are fleshy, ovoid in shape and creamy to yellowish in colour [14]. The species has been recorded in Mozambique, South Africa, Swaziland, Zimbabwe and naturalized in St Helena [1,14]. E. croceum bark is used as an emetic and purgative [15,16] as herbal medicine against human immunodeficiency virus (HIV) opportunistic infections [17]. The bark is also traditionally used to treat and manage tuberculosis [18] and other related diseases such as blood in sputum [18], chest congestion [12,16,[19][20][21], cough [18] and sore throat [16]. The bark is used to manage and treat fever [18] and bark, roots and leaves of the species as herbal medicine to clean the digestive tract [12,16,[19][20][21].  Figure 2). Prinsloo et al [20] isolated a cardiac glucoside, digitoxigenin-glucoside from E. croceum bark extracts ( Figure 2). Yelani et al [21] isolated triterpenoids such as 20-hydroxy-20epitingenone, tingenone, tingenine B, 11αhydroxy-β-amyrin and naringenin from the leaves of E. croceum ( Figure 2).
Prinsloo et al [20] evaluated anti-HIV activities of the compound digitoxigenin-glucoside isolated from E. croceum extracts by assessing the inhibition of recombinant HIV strain in an MT-2 VSV-pseudotyped recombinant virus assay. The compound showed significant inhibition of 90% at 0.2 µM [20]. Prinsloo et al [20,25] evaluated the cytotoxicity of the compound digitoxigeninglucoside using the colourimetric 3-(4,5dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay on Vero cells and scored as percentage reduction at 540 nm of treated culture versus untreated control culture. The isolated compound showed toxicity of only 20% at a concentration of 25 μg/ml on Vero cells with a therapeutic index of 250 [20,25]. Yelani et al [21] evaluated the cytotoxicity of hexane, dichloromethane, and acetone leaf extracts of E. croceum and compounds 20-hydroxy-20epitingenone, tingenone, tingenine B, 11αhydroxy-β-amyrin and naringenin isolated from the leaves of the species using the XTT (sodium 3'-[1-(phenyl amino-carbonyl)-3,4-tetrazolium]bis-[4-methoxy-6-nitro] benzene sulfonic acid hydrate) colorimetric assay against four human cancer cell lines, Vero (monkey kidney), HeLa (human cervix cancer), SNO (human oesophagus cancer) and MCF-7 (human breast cancer) with doxorubicin and zearalenone as positive controls. All extracts showed activities, but significant activities were demonstrated by compounds 20-hydroxy-20-epitingenone and tingenone which showed activities against Vero cells with half maximal inhibitory concentration (IC 50 ) values of 2.7 nM and 8.2 µM, respectively. The cytotoxicity of compounds 20-hydroxy-20-epitingenone and tingenone against three human cancer cell lines, HeLa, MCF-7, and SNO showed IC 50 values ranging between 2.5 µM to 0.4 µM [21].  Gardnerella vaginalis, Neisseria gonorrhoeae and Oligella ureolytica using the broth microdilution method with ciprofloxacin as a positive control. The extract exhibited activities against tested pathogens with MIC values ranging from 1.6 mg/ml to 12.5 mg/ml while ciprofloxacin exhibited MIC value of <0.01 mg/ml [17]. Odeyemi and Afolayan [24] evaluated antibacterial activities of aqueous extract of E. croceum leaves and stem bark against Enterococcus faecalis, Klebsiella pneumoniae, Proteus vulgaris, Salmonella typhimurium, Shigella flexneri and Staphylococcus aureus using the agar well diffusion and broth microdilution methods with amoxicillin (0.0125 mg/mL) as the positive control. The extracts exhibited activities against all the tested pathogens with zone on inhibition ranging from 13.0 mm to 15.7 mm. The control, amoxicillin was active against Enterococcus faecalis, Klebsiella pneumoniae, Shigella flexneri and Staphylococcus aureus with zone on inhibition ranging from 12.5 mm to 26.5 mm. The MIC values exhibited by the extracts and the control against all the pathogens ranged from 0.6 mg/ml to >5.0 mg/ml [24]. The documented antibacterial activities exhibited by E. croceum extracts show the potential of the species as herbal medicine against bacterial pathogens and also corroborate the traditional uses of the species as herbal medicine against sore throat in South Africa [16].

Antifungal activity
Mamba et al [17] evaluated antifungal activities of ethanol extract of E. croceum bark against Candida albicans using the broth microdilution method with ciprofloxacin as a positive control. The extract exhibited activities with MIC value of 1.6 mg/ml while ciprofloxacin exhibited MIC value of <0.01 mg/ml [17]. The documented antifungal activities exhibited by E. croceum extracts show the potential of the species as herbal medicine against fungal and microbial pathogens.  [17]. The documented anti-HIV activities exhibited by E. croceum extracts and the compound digitoxigenin-glucoside isolated from the species corroborate the traditional usage of the species against HIV opportunistic infections in South Africa [17].
Odeyemi and Afolayan [24] evaluated antioxidant activities of acetone extracts of E. croceum leaf and stem bark using the DPPH free radical scavenging, ABTS and FRAP assays with ascorbic acid, butylated hydroxytoluene (BHT) and rutin as reference drugs. The leaf extract IC 50 values for ABTS assay was 0.09 mg/ml, DPPH (0.1 mg/ml) and FRAP (2.5 mg/ml) while bark extract IC 50 values for ABTS assay was 0.2 mg/ml, DPPH (0.07 mg/ml) and FRAP (9.2 mg/ml). The IC 50 values exhibited by the reference drugs, ascorbic acid, BHT and rutin ranged from 0.02 mg/ml to 3.5 mg/ml [24]. The documented antioxidant activities of the extracts of E. croceum are probably a result of flavonoids and phenolics which have been isolated from the species [24,25].

Cytotoxic activity
Prinsloo et al [20,23]  The extract exhibited high cytotoxicity, supporting findings by Yelani et al [21] who argued that cytotoxicity of E. croceum on Vero cells is due to toxic triterpenoids 20-hydroxy-20-epitingenone, tingenone, tingenine B, 11hydroxy-amyrin and naringenin isolated from the leaves of the species.
Despite the long use of E. croceum as herbal medicine in South Africa and Swaziland to treat numerous human diseases, the roots of the species are reported to have been used to cause death by traditional healers [33,34]. Research by Kunene et al [35] showed that E. croceum is categorized as detrimental and causing death of cattle in KwaZulu Natal province, South Africa. Experiments upon rabbits found that 2.5 g per kg body weight, and 10 g of fresh leaves per kg body weight caused death fifteen minutes after administration [36]. Based on these toxicological reports, it can be inferred that E. croceum should be taken with caution when used as herbal medicine.

FINAL REMARKS
Pharmacological studies carried out so far showed that the different extracts and constituents of the species to be promising, which calls for detailed phytochemical, pharmacological and clinical studies of different extracts and chemical constituents of the species. Determination of the active principles is very important in the extracts and compounds of the species in order to avoid life-threatening complications as the species is widely used as herbal medicine. There is lack of information about the effect of prolonged consumption of E. croceum extracts. Therefore, future studies should include the identification of toxic compounds, possible side effects caused by taking E. croceum as herbal medicine, and mechanisms of how potential toxic components of the species can be managed.