Submitted in partial fulfillment of the requirements for the Degree of Master of Technology: Biological Sciences, Technikon Natal, 2000. / This study focussed on the anti-bacterial, anti-fungal and anti-protozoal activity of four plant extracts, maackiain and formononetin from Virgilia oroboides and chlorophorin and lroko from Chlorophora excelsa / M

Schumanniophyton magnificum and Glypheae brevis are important medicinal plants growing wild in the West African rain forest. They are used in folkloric medicine in the treatment of epilepsy and convulsion as well as for some other diseases. The purpose of this work was to investigate the aspect of folkloric use in order to support folkloric claims and document the findings. The extracts were prepared from ground plant material by a continuous extraction method. Five hundred grams of ground plant material were continuously de-fatted with 2 L petroleum ether (60°- 80°) in a Soxhlet apparatus for about 5 h. The resulting marc was dried and the chemical constituents extracted hot in a Soxhlet apparatus for about 8 to 10 h with 2 L aqueous ethanol (70%). The efficacy of the extraction method was confirmed using standard bioassays and phytochemical analyses. The anti-convulsant activity of the crude extracts was evaluated in vivo against chemically induced convulsions using three different animal models, namely the strychnine, the picrotoxin and the pentylenetetrazole tests. The acute and delayed toxicity test results showed that in all the animal models investigated very high doses, about four times higher than the protective doses of the extracts, were required to kill 50% of the population of animal used. Phytochemical assays of the extracts indicated the presence of alkaloids only in S. magnificum root extract and glycosides in extracts from both species. The glycosides were positive to Baljet, Xanthydrol and Keller-Kiliani tests for cardiac glycosides. S. magnificum and G. brevis chemical constituents were initially isolated with a sequential fractionation method starting with a highly non-polar solvent and gradually increasing to a more polar solvent. The fractions were pooled on the basis of TLC similarity profiles when viewed under the UV light at 254 and 366 nm and were found to have two and four major UV absorbing fractions for S. magnificum and G. brevis respectively. Radio-receptor binding tests were used to assess the anti-convulsant activities of the hydro-alcoholic crude extracts, the organic and aqueous fractions of the crude extracts, partially purified components and pure components in in vitro tests against some standard GABA[A] receptor antagonists, muscimol and isoguvacine respectively. The anti-convulsant activities resided in the aqueous fractions of the hydro-alcoholic crude extracts of both plants. The purely organic fractions of G. brevis demonstrated no activity while all the fractions of the aqueous component demonstrated some degree of activity. The anti-convulsant activity of S. magnificum was found only in one fraction-Fraction 1. This Fraction was further investigated and one of the components appear to be responsible for the activity. The structure of the active constituent was 5,7dihdroxy-2 methylbenzopyran-4-one, a noreugenin. A second bioactive compound, schumanniofoside, was identified from Fraction M[5.2] from S. magnificum. / Thesis (Ph.D.)-University of Natal, Pietermaritzburg, 2002.

The drug aloes has been known since earliest times and is mentioned in the Ebers papyrus of circa 1,500 B.C. Alexander the Great is reported to have sent a commission to Socotra to investigate the aloes grown there. The chemical composition of aloes is complex, and being of plant origin, subject to variation. Both the complexity of the chemical constituents and their biological variation has resulted in a very large volume of conflicting material being published on this drug export. Since aloes is used as a purgative for both human and veterinary use, it is obviously important that the dosage and consequently the active constituents, should comply to an accurate means of standardisation. To date, despite extensive world wide research into this drug such standardisation has not been achieved. Even the methods used for the assay of the principal constituent, aloin, vary considerably in their results, and to complicate matters new chemical principles have been isolated from aloes in recent years. Consequently the purpose of this work has been to investigate the main chemical assay methods currently in use, and to determine which was the most accurate, and why discrepancies occurred in the selected assay methods. furthermore the results obtained by chemical assay have been compared with those obtained by biological assay in an attempt to correlate aloin content with purgative activity. Newly isolated compounds have been investigated biologically for the first time, and the biological assays of the resinous, glycosidal and other compounds of aloes have been performed. Intro. p.1-2.

Introduction: Despite the tremendous advances made with regard to synthetic organic medicinals within the last two decades, heavy reliance is still placed on plant products. This is especially true of the anthracene derivatives used medicinally as purgatives, and which are derived principally from senna, cascara, rhubarb, frangula and aloes. While particular attention has been paid to the chemistry of the former group in recent years, aloes has been largely neglected, possibly due to the fact that the Aloe species are confined largely to areas where extensive research facilities are lacking, such as Africa , India and the West Indies. Thus research in Europe has been confined largely to the lump aloes of commerce, derived from relatively few species. In 1953 a comprehensive report by Hodge (103) appeared on "The Drug Aloes of Commerce, with Special Reference to the Cape Species". Hodge observed that South Africa abounds in species just as abundant as A.ferox, (which is the prime source of Cape aloes), and advised that a systematic chemical survey might show certain of these to be not only higher yielders of bitter aloetic juice but also sources of a superior drug product. Consequently an investigation along these lines is presented here, and it is observed that several species apart from A.ferox not only contain aloin, but also yield a large volume of aloetic juice. Only pharmacologic studies can reveal if the juice of these species is as safe as that of A.ferox, but without doubt they could be used for the extraction of crystalline aloin. Concurrently, the distribution of the Aloe resins, said by some to be purgative themselves, has been studied. The investigation has revealed that the structurally similar compound homonataloin enjoys an equally wide distribution as aloin. However, almost invariably it is confined to small species yielding little aloetic juice, apart from which nothing is known regarding its pharmacologic properties. It is interesting to note that the resin distribution in the homonataloin-containing species is very similar to that of the aloin-containing species, but differs widely from. that of the species containing neither of these principles. Apart from aloin and homonataloin, aloinoside and chrysophanol also occur in Aloe species, and together with the resins, these indicate that when all the South African Aloe species have been investigated, they may well be of chemotaxonomic value. Within the comparatively short space of the last decade some work has been performed on aspects of the metabolism of such anthracene-containing species as Rheum, Rhamnus and Rumex. These investigations have shown that the anthracene derivatives are not merely waste products, but perform definite metabolic functions. The latter portion of this work has been devoted to this relatively neglected aspect of the Aloe species.

Aeromonas hydrophila is distributed widely in nature and is responsible for an array of human infections. Several studies on the isolation and characterisation of the organism abound. Although there are reports on the antibiotic resistance profiles of the organism, these reports have not been updated in Limpopo province, South Africa despite the established fact that antibiograms vary with time and geographical area. Antibiotic resistance and pathogenesis of an organism are dependent on a host of factors such as the production of extended spectrum beta-lactamases and the genetic profiles such as the genes coding for resistance and possession of integrons and how these characteristics overach with the phylogenetic inter-relatedness of isolates from different sources. In spite of the aforementioned concerns on the efficacy of antibiotics due to the acquisition or endowment of microorganisms with intrinsic and extrinsic factors , which enhances resistance to antibiotics , medicinal plants are reportedly offering promise as alternative sources of efficacious management of infections. Medicinal plants are employed by traditional healers in the management of infections in developing countries especially in Africa. However, the antimicrobial activities of medicinal plants against Aeromonas hydrophila have received only a cursory attention. In an endeavour to undertake a comprehensive study on the isolation, characterisation, antibiograms, activities of medicinal plants as well as the genetic profiles, including phylogenetics relatedness of Aeromonas isolates from different sources, stool and water samples were collected over a two year period from designated places in Limpopo Province and analysed using standard techniques applicable to the constituent research activity. The research findings are presented in six chapters as presented hereunder. The first chapter focussed on the literature review of the organism and reflects areas such as the morphology, laboratory diagnosis, clinical manifestations, pathogenesis, antimicrobial susceptibility profiles, antibacterial activities of medicinal plants as well as the genetic aspects of Aeromonas hydrophila. / Environmental Science / D.Phil. (Environmental Science)

The post-harvest physiology of nine frequently used indigenous southern African medicinal plants was investigated, in particular the effects of storage time and accelerated ageing on the biological activity and chemical constituents of these plants. Water, ethanol and hexane extracts of fresh plant material as well as material that had been stored in dry form in paper bags at room temperature for 90 days (short-term) were tested. Three bioassays, the COX-1 anti-inflammatory assay, nematode anthelmintic assay and minimum inhibitory concentration anti-bacterial assay, were used to determine biological activity. Thin layer chromatography of all the plant extracts were used to determine changes in chemical composition. The plants tested were Alepidea amatymbica Eckl. & Zeyh., Leonotis leonurus (L.) R. Br., Drimia robusta Bak., Vernonia colorata (Willd.) Drake, Scilla natalensis Planch., Eucomis autumnalis (Mill.) Chitt. subsp. autumnalis, Bowiea volubilis Harv. ex Hook. f., Helichrysum cymosum (L.) D. Don and Siphonochilus aethiopicus (Schweinf.) B. L. Burtt. Only those plants, which are known to exhibit a particular biological activity either traditionally or scientifically, were tested in the relevant bioassays. Of the plant extracts tested for anthelmintic activity only the water extracts showed activity and very little change in activity was observed after storage. Of the plant extracts tested for anti-inflammatory activity the ethanol extracts generally yielded highest activity. S. natalensis and B. volubilis both showed an increase in cyclooxygenase inhibition (anti-inflammatory) activity after storage whereas S. aethiopicus, H. cymosum, D. robusta and V. colorata showed a loss in activity after storage. The anti-inflammatory activity of E. autumnalis did not change. The water extracts of plants tested for antibacterial activity showed no activity, whereas the ethanol extracts generally showed an increase in activity. The TLC fingerprints indicated that there was chemical break-down during storage in certain species. These corresponded to the changes in biological activity. Alepidea amatymbica, Eucomis autumnalis, Helichrysum cymosum, Leonotis leonurus, Siphonochilus aethiopicus and Vernonia colorata were investigated further as to the effect of one year's storage (long-term storage) on their chemical composition and biological activity. Similar trends to that of the 90-day storage were observed. Activity gained in plants that were stored for 90 days was retained after a year of storage. Elevated temperature and humidity (55 C and 100% relative humidity) were used to accelerate the ageing process of Alepidea amatymbica, Leonotis leonurus and Vernonia colorata plant material. Again changes in the chemical composition and biological activity were observed, and the extent of these changes was greater than those in the stored material. The compounds responsible for the cyclooxygenase inhibition in the ethanolic extracts of Alepidea amatymbica leaf material appear to be stable and were not affected by the conditions of the accelerated ageing procedure (55 C and 100% humidity for seven days), but the root material lost activity, as did the leaf material of Leonotis leonurus. The leaf material of Vernonia colorata showed a slight (8%) increase in cyclooxygenase inhibition activity. The response of the plant material to accelerated ageing with respect to antibacterial activity varied with plant species. Alepidea amatymbica root material and Vernonia colorata leaf material appear to be stable whereas the other plant materials lost activity after prolonged (25 days) ageing. / Thesis (M.Sc.)-University of Natal, Pietermaritzburg, 2003.

Aeromonas hydrophila is distributed widely in nature and is responsible for an array of human infections. Several studies on the isolation and characterisation of the organism abound. Although there are reports on the antibiotic resistance profiles of the organism, these reports have not been updated in Limpopo province, South Africa despite the established fact that antibiograms vary with time and geographical area. Antibiotic resistance and pathogenesis of an organism are dependent on a host of factors such as the production of extended spectrum beta-lactamases and the genetic profiles such as the genes coding for resistance and possession of integrons and how these characteristics overach with the phylogenetic inter-relatedness of isolates from different sources. In spite of the aforementioned concerns on the efficacy of antibiotics due to the acquisition or endowment of microorganisms with intrinsic and extrinsic factors , which enhances resistance to antibiotics , medicinal plants are reportedly offering promise as alternative sources of efficacious management of infections. Medicinal plants are employed by traditional healers in the management of infections in developing countries especially in Africa. However, the antimicrobial activities of medicinal plants against Aeromonas hydrophila have received only a cursory attention. In an endeavour to undertake a comprehensive study on the isolation, characterisation, antibiograms, activities of medicinal plants as well as the genetic profiles, including phylogenetics relatedness of Aeromonas isolates from different sources, stool and water samples were collected over a two year period from designated places in Limpopo Province and analysed using standard techniques applicable to the constituent research activity. The research findings are presented in six chapters as presented hereunder. The first chapter focussed on the literature review of the organism and reflects areas such as the morphology, laboratory diagnosis, clinical manifestations, pathogenesis, antimicrobial susceptibility profiles, antibacterial activities of medicinal plants as well as the genetic aspects of Aeromonas hydrophila. / Environmental Science / D.Phil. (Environmental Science)