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Traditional herbal drugs of southern Uganda Part III: Isolation and methods for physical characterization of bioactive alkanols from Rubus apetalus

Authors:
  • IHRC / FDA Office of Laboratory Safety

Abstract

The East African plant Rubus apetalus Poir. was collected as a component of an ethnobotanical survey in southern Uganda. No phytochemical investigations of this plant have been found in the literature. Preliminary antimicrobial susceptibility tests performed in Uganda indicated biological activity against several bacterial and one fungal human pathogen. Bulk re-collection of Rubus apetalus was accomplished and crude extraction performed in preparation for further testing. Two chemical fractions of the crude extract were active in the antimicrobial susceptibility assay. Fractionation of one of the active crude fractions led to the isolation and elucidation of a mixture of related compounds that exhibit antimicrobial activity against Staphylococcus aureus (MIC=62 microg/ml), Streptococcus faecalis (16 microg/ml) and Candida albicans (32 microg/ml).
... Rubus apetalus, one of such plants, is an annual scrambling shrub from the Rosaceae family. This tropical plant has a cluster of stems from a woody rootstock and can grow to around 150 cm tall [20]. Its fruits are used as a local food, though the plant is sometimes cultivated in gardens [21,22]. ...
... A decoction of the ripe fruit is mixed with sugar and used to treat anemia while an infusion of the leaves is used for the management of diabetes mellitus [23]. R. apetalus is also used as fertility booster [24], anti-venom [25], anti-fungi [20], and anti-bacteria [26]. Moreover, the in vitro [27,28] and in vivo antioxidant potentials of R. Apetalus have been reported and could be justified by its contents in bioactive compounds such as saponins, alkaloids, flavonoids, tannins, triterpenoids and vitamins [23]. ...
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Background: Cryptorchidism (CPT) is an important cause of male infertility. Rubus apetalus is a medicinal plant with a powerful antioxidant potential. We investigated the effects of aqueous and methanolic extracts of R. apetalus on spermatozoa parameters, antioxidant enzymes and fertility potential of rats with experimental unilateral CPT. Method: Normal (n = 15), sham-operated (n = 15) and cryptorchid rats (n = 80; distributed into 16 groups of 5 rats/group) were treated for 2, 4 or 8 weeks with either distilled water (10 ml/kg/day), vitamin E (75 mg/kg/day), aqueous or methanolic extract of R. apetalus (12 and 60 mg/kg). Sex organ weights, spermatozoa parameters, testicular proteins, sex hormones, fertility potential, morphometric characteristics of testis and oxidative stress markers were measured. Results: CPT significantly (p < 0.05-0.001) decreased testicular and epididymal weights, spermatozoa density, spermatozoa motility, spermatozoa normality, testicular proteins, LH, FSH and testosterone concentrations. In cryptorchid rats, peri-vascular fibrosis significantly increased (p < 0.001), while diameter of the seminiferous tube, germ cell thickness, gestation index and fertility index decreased when compared to control. Additionally, CPT induced oxidative stress by increasing lipid peroxidation and by reducing superoxide dismutase and catalase activities. These alterations were corrected by R. apetalus. For instance, a significantly increase (p < 0.05-0.001) in spermatozoa motility, normality, viability and density after 2, 4 and 8 weeks of treatment was noticed. R. apetalus also increased (p < 0.05-0.001) testicular proteins, gestation index (90-100%) and fertility index (90-100%), compared to the untreated cryptorchid rats. Conclusion: R. apetalus boosts fertility potential in cryptorchid rats and could be considered as a promising alternative agent for the management of infertility associated with CPT.
... Spectra were recorded in deuterated chloroform (CDCl3) using Tetramethylsilane (TMS) as the internal standard for 1H NMR [18] (TMS at 0 ppm, CDCl3 at 7.26 ppm) 13 C NMR (CDCl3 at 77.0 ppm) and DEPT [19] on a Bruker Avance spectrometer (Bruker, Switzerland). The DEPT experiments were used to determine the multiplicities of carbon atoms. ...
... The remaining peaks include 32.76 (C-2), 31.88 (C-4), 29.66-29.32 (C-6-C(n-1)), 25.69 (C3), 22.66 (C5).The proton and carbon assignments were made based on DEPT 135 and previously reported data [19] (Supplementary Fig. 1C). EI-MS spectrum of isolate A lacked single molecular ion and more than one [M-H 2 O] + peaks were observed suggesting it to be a mixture of alkanols, varying slightly in their chain length ( Supplementary Fig. 1D). ...
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Background: Chromolaena odorata, has been traditionally known for its insect repellent property. Aim of this study was to determine larvicidal tendency of C. odorata on Culex quinquefasciatus and isolate compounds responsible for this activity and to determine the mechanism of action of these compounds. Methods: C. odorata plant extract was screened for mosquito larvicidal activity. The extract was fractionated using chromatography and the bioactive fraction showing larvicidal activity was identified. The chemical nature of the compounds in the bioactive fraction was determined using NMR and Mass spectrometry. Results: We identified phytosterols and alkanols to be the compounds regulating larvicidal activity in the bioactive fraction of the plant extract. Stigmasterol and 1-hexacosanol were identified to be the chief orchestrators of larvicidal activity and their mode of action has been observed to be neurotoxicity. At a molecular level both stigmasterol and 1-hexacosanol were found to be inhibiting acetylcholinesterase activity in C. quinquefasciatus & A. aegypti. The acetylcholinesterase inhibitory effect was validated in vitro using recombinant acetylcholinesterase and ex vivo in larval homogenates of Culex and Aedes. Electrophysiological studies using electroantennography have shown enhanced neural response to these compounds. Conclusions: Neurotoxic effect of C. odorata derived stigmasterol and 1-hexacosanol, exerted through acetylcholinesterase inhibition was responsible for the mortality of C. quinquefasciatus, A. aegypti &Chironomus riparius. EAG studies pointed out hyper-excitability of the olfactory system by these compounds. General significance: These compounds are natural agents for mosquito control that can be used in vector control as larvicidal compounds, pending further investigations.
... Also reported medicinal properties of V. amygdalina include antioxidant activity [13], [14], anticancer/tumor properties [15], [16], [17] [18], hepatoprotective and nephron -protective properties [19], oxytocic property [20] [21] and pesticidal property [22], [23]. Ethno-botanically, the leaves are particularly utilized in the treatment of malaria, diabetes mellitus, venereal diseases, wounds, hepatitis and cancer [24], [25], [26], [27]. Colletotrichum gloeosporioides is a pathogenic fungus of the Cashew tree, but it was also found as an endophyte in many cases [28]. ...
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The emergence of antibiotic-resistant strains of microorganisms calls for research and the development of strategies to curb this menace. The screening of antimicrobial compounds from endophytic fungi is a promising way to meet the increasing threat of drug-resistant. This study aims at evaluating the antimicrobial activity of the endophytic fungal extract of Colletotrichum sp. from the leaves of Vernonia amygdalina (Del). Endophytic fungal isolation, cultivation, and identification were carried out using standard methods. Subsequently, fermentation and extraction of the secondary metabolites were done on a rice medium. About 1.2g of the fungal extract was subjected to vacuum liquid chromatography (VLC) using gradient-based elution; n-hexane and ethyl acetate followed by dichloromethane and methanol. The high-performance liquid chromatography (HPLC) analysis of the crude extract and VLC fractions were also carried out. Gel permeation chromatography using Sephadex LH-20 was carried out on Fraction JT2. Antimicrobial Assays for fraction JT4 and the Sephadex LH-20 fractions (JT2C-JT2E) were also carried out. The results were analyzed using Statistical Package for Social Sciences. The high-performance liquid chromatography (HPLC) analysis of the crude extract and VLC fractions revealed the presence of the five bioactive metabolites (acropyrone, beauvericin, indole-3-carbaldehyde, indolyl-3-acetic acid, and rocaglamid A). The result of the antimicrobial activity of the VLC fraction, JT4, showed that the fraction exhibited a significant inhibitory effect against Bacillus subtilis, Pseudomonas aeruginosa at 0.25-1 mg/ml (IZD) and antifungal activity against Aspergillus niger only at 0.0625-1 mg/ml (IZD). The result of the antimicrobial activity of the gel chromatographic fractions revealed a significant effect against Staphylococcus aureus, Pseudomonas aeruginosa and Escherichia coli at 0.0625-1 mg/ml and had no inhibitory effect against Bacillus subtilis, Aspergillus niger and Candida albicans. The results of this study show that endophytic fungi on V. amygdalina could be a potential source of bioactive compounds for the development of potential and effective antibiotics.
... The leaves are particularly utilized in the treatment of malaria, diabetes mellitus, venereal diseases, wounds, hepatitis and cancer [11,12,13]. The anthelminthic properties of the plants amongst the diverse uses of the plant in combating diseases have also been reported [14]. ...
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The endophytic fungus, colletrotricum species, isolated from the leaves of Vernonia amygdalina was investigated for its chemical constituents and biological activity. The pure fungus was grown using solid fermentation on rice medium and the metabolites were extracted using ethyl acetate. Further purification of the extract was carried out using vacuum liquid chromatography (VLC) and gel chromatography on Sephadex LH-20. The chemical constituents were detected by dereplication using HPLC-DAD. Antimicrobial assay of the Sephadex fractions EC3-1 and EC3-3 were evaluated using agar well diffusion assay against seven pathogenic microorganisms (Staphylococcus aureus, Escherichia coli, Salmonella typhi, Bacillus subtilis, Pseudomonas aeruginosa, Aspergillus niger and Candida albicans). EC3-3 was also subjected to antioxidant assay using DPPH free radical scavenging model. HPLC-DAD analysis of the VLC fractions revealed the presence of compounds like Hydroxybenzaldehyde, Cladosporin, Desmethyldichlorodiaportin and p-Hydroxybenzoic acid. EC3-1 and EC3-3 showed mild antimicrobial activity against S. aureus, S. typhi and A. niger strains, with minimal inhibitory concentrations ranging from 0.19 to 0.46 mg/mL. EC3-3 was found to exhibit very high ability to scavenge DPPH radicals with IC50 of 3.16 µg/mL compared to Ascorbic acid (IC50 of 50.72 µg/mL). HPLC analysis of EC3-3 revealed the presence of Palitantin, which has been shown to possess antimicrobial and antioxidant activity, and two unidentified major peaks. The endophytic fungus, Colletotrichum species isolated from Vernonia amydalina produced bioactive metabolites which exhibited antimicrobial and antioxidant activities thus projecting Vernonia amydalina as a potential source of fungal endophytes that can be further studied for generation of novel bioactive compounds.
... Vernonia amygdalina, a member of Compositae family, is commonly known as bitterleaf. It possess characteristic astringent bitter taste and the leaves are employed as vegetable in soups, in treatment of malaria, diabetes mellitus, venereal disease, wounds, hepatitis and cancer (Kambizi and Afolayan, 2001;Hamill et al., 2003;Erasto et al., 2007). Jatropha tanjorensis is a perennial herb, a member of the Euphorbiaceae family, commonly called 'hospital too far', catholic vegetable, Jatropha. ...
... The East African plant, R. apetalus was collected as a component of an ethnobotanical survey in southern Uganda. Preliminary antimicrobial susceptibility tests performed indicated the biological activity of the plant against several bacterial and fungal human pathogens, such as Staphylococcus aureus, Streptococcus faecalis and Candida albicans [77]. Examples of other antimicrobial Rubus extracts included R. ulmifolius, a Chilean folk medicine [78]. ...
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Species of the genus Rubus (family Rosaceae, sub-family Rosoideae, tribe Potentilleae) are widely distributed in the warmer temperate zones of the northern hemisphere. Rubus species have been known since ancient times for their curative properties, and have been used for the treatment of various ailments, such as wounds, diarrhea, colic pain, diabetes, inflammatory disorders, and as antimicrobial agents. However, their use has been based mainly on empirical findings. This contribution provides a comprehensive review of the work published on the phytochemistry and pharmacology of the Rubus species over the past few years, in order to obtain a better understanding of the biological significance of this large genus.
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Background. Many studies have been undertaken on the medicinal values of Erythrina abyssinica Lam. ex DC. (Fabaceae). The details, however, are highly fragmented in different journals, libraries, and other publication media. This study was therefore conducted to provide a comprehensive report on its ethnobotany, ethnomedicinal uses, phytochemicals, and the available pharmacological evidence supporting its efficacy and safety in traditional medicine. Method. We collected data using a PROSPERO registered systematic review protocol on the ethnobotany, phytochemistry, and ethnopharmacology of Erythrina abyssinica from 132 reports that were retrieved from electronic databases. Documented local names, morphology, growth habit and habitat, ethnomedicinal and nonmedicinal uses, diseases treated, parts used, method of preparation and administration, extraction and chemical identity of isolated compounds, and efficacy and toxicity of extracts and isolated compounds were captured. Numerical data were summarized into means, percentages, and frequencies and presented as graphs and tables. Results. Erythrina abyssinica is harvested by traditional herbal medicine practitioners in East, Central, and South African communities to prepare herbal remedies for various human and livestock ailments. These include bacterial and fungal infections, tuberculosis, malaria, HIV/AIDS, diarrhea, cancer, meningitis, inflammatory diseases, urinary tract infections, wounds, diabetes mellitus, and skin and soft tissue injuries. Different extracts and phytochemicals from parts of E. abyssinica have been scientifically proven to possess anti-inflammatory, antibacterial, antioxidant, antiplasmodial, antiproliferative, antifungal, antimycobacterial, antidiarrheal, anti-HIV 1, antidiabetic, and antiobesity activities. This versatile pharmacological activity is due to the abundant flavonoids, alkaloids, and terpenoids present in its different parts. Conclusion. Erythrina abyssinica is an important ethnomedicinal plant in Africa harboring useful pharmacologically active phytochemicals against various diseases with significant efficacies and minimal toxicity to mammalian cells. Therefore, this plant should be conserved and its potential to provide novel molecules against diseases be explored further. Clinical trials that evaluate the efficacy and safety of extracts and isolated compounds from E. abyssinica are recommended. 1. Introduction Erythrina abyssinica Lam. ex DC. (Fabaceae) is an important medicinal plant as evidenced by the existence of its names in various local languages and high frequency of citation in ethnobotanical surveys [1–4]. The genus Erythrina derives from the Greek word “erythros,” translated to mean red (a reflection of the showy red flowers of its various species). The epithet ‘‘abyssinica’’ means ‘‘from Ethiopia’’ [5]. The Erythrina genus houses at least 120 species distributed mainly in tropical and subtropical zones [6]. Plants in this genus are usually referred to as “coral trees” due to their red flowers and branches that resemble the shape of sea coral [7]. Erythrina abyssinica is a deciduous leguminous tree native to East Africa but also found in Central and South Africa [8, 9]. Tropical Asia and Central America have E. abyssinica as an exotic species. The common English names of E. abyssinica are coral tree, Uganda coral, kaffir boom, erythrina, flame tree, red-hot-poker tree, and lucky-bean tree [10]. Some of the local names used across indigenous communities are summarized in Table 1. Folk name (local language) Country Authors Ejjirikiti (Luganda), Murinzi, Kiko Omoko/Echuko (Rutoro, Rukonzo), Oluo (Lugbara), Kisoro, Lochoro, Oding, Loting (Acholi), Kikiri (Kwamba), Engosorot (Ateso), Olawu (Madi), Koli (Jopadhola), Owila kot (Lango), Muyirikiti, Ekilama (Lusoga), Cheroguru, Muragolo (Lugishu), Mutembetembe (Lugwe), Bwiko (Lukiga), Kaborte (Sebei), Kiko, Muko (Lunyangkore, Lutoro), Mudongodongo, Mukobe (Lunyuli) Uganda [2, 3, 10–15] Omotembe (Kisii), Muhuti (Kikuyu), Ekirikiti or Ol-Goroshe (Maasai), Muuti (Meru), Kivuti or Muvuti (Kamba), Mulungu (Taita), Mwamba ngoma, Mbamba ngoma, Muhuti, Mjafari or Mwamba (Kiswahili), Kumurembei (Luhya) Kenya [10, 16–19] Qanqari (Iraqw), Mriri (Chagga), Muhemi (Hehe), and Muungu (Pare), Kisebhe (Rungwe) Tanzania [20–22] Kuara, Korra, Korch (Amharic) Ethiopia [10] Umuko (Lunyarwanda) Rwanda [23–26] Dus (Arabic), Hab al Arous Sudan, South Sudan [10, 27, 28] Chisunga (Lunda) Democratic Republic of Congo [10] Mulunku (Chokwe) Angola [4] Mulunguti, Mwale (Nyanja) Mozambique, Zimbabwe, Zambia, Malawi [10] Mulunguti (Bemba, Tongan) Zambia, Mozambique, Zimbabwe [5, 10] Mutiti (Shona) Zimbabwe [5] Suwawue, Soaueh (Tigrigna) Eritrea, Ethiopia [10, 29]
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Background There is high mortality and morbidity due to poisonous snakebites globally, with Sub Saharan African having one of the highest rates. However, Traditional Medicine Practitioners (TMP) have been treating snakebites in Uganda for long. However, few studies have been conducted to document such vital traditional indigenous knowledge before its lost. The aim of this study was to document the medicinal plant species used by experienced TMP in treating snake envenomation in selected post-conflict parts of northern Uganda. Methods An ethnopharmacological survey was conducted in Serere, Kaberamaido and Kaabong districts in Uganda. Twenty-five TMP with expertise in treating snakebites were purposively identified using the snowball technique, and interviewed using semi structured questionnaires. Data were analysed using simple descriptive statistics. Results Sixty plant species from 28 families were documented with high consensus among the isolated Ik community in Kaabong district. Most of the plant species used belonged to the Asteraceae and Fabaceae families with eight species each. Additionally, the genus Echinops was the most well represented with three species. The most commonly used plant species by frequency of citation were: Steganotaenia araliaceae (16), Microglossa pyrifolia (Lam.) and Gladiolus dalenii Van Geel (13), Aframomum mildbraedii Loes. (11), Jasminum schimperi Vatke, Cyathula uncinulata (Schrad) Schinz (10), Crinum macowanii Baker and Cyphostemma cyphopetalum (Fresen.) Desc. ex Wild & R.B.Drumm (10). S. araliaceae which was mentioned by all the TMP in the Ik community was used as first aid. Most of the plant species were harvested from the wild (68.75%) and were herbs (65.0%) and trees (23.3%). The most commonly used plant parts were roots (42.6%) and leaves (25.0%). Thirteen different methods of preparation and administration were used. Most of herbs were administered orally (61.2%), and topically (37.6%). The commonest methods of oral application were cold water infusions (32.5%) and decoctions (21.7%). Conclusions TMP widely use several medicinal plant species for treating snakebites envenomation in the selected post-conflict sub-regions of Acholi, Teso and Karamoja in Uganda
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