The medicinal chemistry of selected marine algae indigenous to South Africa was investigated. Following the isolation and characterisation of a number of new and known compounds, the associated in vitro cytotoxic profiles of these new compounds was investigated. Plocamium maxillosum yielded two new cyclic polyhalogenated monoterpenes which were characterised as 2E-chloromethine-4E-chlorovinyl-4-methyl-5-cyclohexen-1-one (2.1) and 2Z-chloromethine-4E-chlorovinyl-4-methyl-5-cyclohexen-1-one (2.2) on the basis of one and two dimensional NMR spectroscopic data and MS analysis. These compounds were also found to have good cytotoxic activity against breast cancer cell lines. Although these compounds are based on a regular monoterpene skeleton, they represent an uncommon feature not often seen in cyclic halogenated monoterpenes from marine algae. Plocamium robertiae yielded one new cyclic polyhalogenated monoterpene identified as 4,5- dibromo-5-chloromethyl-1-chlorovinyl-2-chloro-methylcyclohexane (2.6) and one known compound called 2,4-dichloro-1-chlorovinyl-1-methylcyclohexane-5-ene or Plocamene D (2.9). Portieria hornemannii was collected from Port Edward in Natal and yielded three new compounds, namely; 3Z-1,6-dibromo-3-(bromomethylidene)-2,7-dichloro-7-methyloctane (3.1), 1E,3Z-1,6-dibromo-3-(bromomethylidene)-7-chloro-7-methyloct-1-ene (3.2), 1Z,3Z- 1,6-dibromo-3-(bromomethylidene)-7-chloro-7-methyloct-1-ene (3.3), and one known compound, namely; 3S,6R-6-bromo-3-(bromomethyl)-3,7-dichloro-7-methyloct-1-ene (3.4). Compounds 3.1 and 3.2 showed no cytotoxic activity against breast cancer cells. Another Portieria hornemannii sample was collected from Noordhoek in the Eastern Cape, it yielded one known compound referred to as 3Z-6-bromo-3-(bromomethylidene)-2,7- dichloro-7-methyloct-1-ene (3.5), as well as one new compound called portieric acid A (3.6) or 5-bromo-2-(bromomethylidene)-6-chloro-6-methylheptanoic acid. Portieric acid A showed slight cytotoxic activity and also represents a new class of compound within the genus Portieria. The isolation of secondary metabolites from the South African red alga, Laurencia glomerata, yielded two known compounds; 7-hydroxylaurene (4.9) and cis-neolaurencenyne (4.12), as well as one chamigrane related compound (4.11). Laurencia flexuosa yielded one known compound called 3Z-bromofucin (4.13). Using 1H NMR, GC and molecular systematics, a novel method for identifying different species of Laurencia was also investigated.

In studies on the bioactivities of selected eastern Cape seaweeds, a suitable extraction solvent, namely methanol was used. The antimicrobial, antineoplastic and antiviral activities were investigated, and a seasonal comparison of antimicrobial activities as well as a scanning electron microscopic study were included. A number of algae were found to display significantly high antibacterial activities, of these, the two red algae Plocamium corallorhiza and Laurencia glomerata and the two brown algae Zonaria subarticulata and Ecklonia biruncinata showed the most potent broad spectrum antibacterial activity. L.glomerata and E.biruncinata were active against all five test bacteria, but were inactive against the yeast Candida albicans. C.albicans was the most resistant organism tested,~with only Peyssonelia capensis, f-corallorhiza and Ulva rigida inhibiting its growth. Of the 17 seaweeds tested, none showed any clear seasonal changes in antimicrobial activity. The microorganisms however did vary in their susceptibility to treatment. Staphylococcus aureus and the Micrococcus species were the most susceptible to treatment by the algal extracts, with the Gram positive endospore former, Bacillus subtilis and the two Gram negative bacteria Escherichia coli and Pseudomonas aeruginosa showing more resistance to treatment. C.albicans however was the most resistant organism. In the antineoplastic studies, the brown algae Z.subarticulata, E.biruncinata and Sargassum heterophyllum showed potent activity against both the normal, LLCMK, and cancerous, mouse melanoma-3S0 cells, reducing cell growth in each case. The green algae showed potent activity against the cancerous cells, lowering growth to 30% that of the normal cells. Normal cell growth was unaffected or was stimulated by the presence of these algal extracts. The red algae showed no antineoplastic activity. Representatives of each of the red, brown and green algae were used in the brine shrimp (Artemia salina) cytotoxicity study. None of the algae showed any toxicity towards the brine shrimp. In the antiviral studies against Polio Type 1, strain L-Sa, a reduction in virus infectivity was used as an indication of the presence of antiviral properties in the seaweeds tested. This was done by comparing the virus titres of algal-treated cells with those of untreated cells. Six of the algae tested displayed antiviral activity, these were the two Rhodophyceae Hypnea spicifera and L.glomerata, the two Phaeophyceae Dictyopteris macrocarpa and Dictyota naevosa, and the two Chlorophyceae U.rigida and Halimeda cuneata. Of these, D.naevosa showed particularly strong activity, recording a log reduction in virus titre of 4.

During 1982 benthic and floating filamentous algal mats appeared in the maturation ponds of the Grahamstown Sewage Disposal Works. These mats clogged the ponds and reduced the efficiency by which the effluent was purified. As they continued to be a problem despite numerous efforts to remove them, this study was initiated to investigate the alga, establish why it was successful in the pond environment, and how its growth could be controlled. The physico-chemical environment of a representative maturation pond was characterised while laboratory studies were conducted to investigate the growth, photosynthetic and respiratory characteristics in the alga. The alga was identified as Rhizoclonium riparium (Roth) Harv. although its morphological variability was greater than that reported in the literature. Growth and photosynthetic studies indicated that the alga favoured temperatures between 20 and 30⁰C and relatively high light intensity (700μE.m⁻².s⁻¹) and that it was highly productive. In addition, the alga exhibited photoadaptive ability, although it seemed to be sensitive to photoinhibition. Its success in the maturation pond was attributed to the favourable physico-chemical environment, particularly the high transparency of the effluent which allowed the penetration of PAR to the pond floor and to the alga's ability to adapt to the change in environment when it floated from the pond floor to its surface. Although the algal mats contribute to the oxidation of the effuent within the maturation pond, they are generally detrimental to the system because they shade the water column and inhibit wind-induced mixing. Recommendations on methods of controlling of the mats are presented. / KMBT_363 / Adobe Acrobat 9.53 Paper Capture Plug-in

Southern African biogeographic boundaries delimit the phylogeographic distribution of some coastal and estuarine invertebrates. This study investigated the impact of these boundaries on the phylogeographic distribution of two intertidal red seaweeds, Gelidium pristoides and Hypnea spicifera using the mitochondrial Cox2-3 spacer and the nuclear ITS1 regions. G. pristoides spores have short distance-dispersal, while long distance-dispersal is more likely in H. spicifera via spores and drifting fertile thallus fragments. Both markers revealed a south-western and south-eastern lineage within G. pristoides but the breaks between lineages do not coincide with any recognised biogeographic limits. The Cox2-3 spacer revealed a boundary between the two lineages at the Alexandria Coastal Dunefield (ACD) and ITS1 at the Gamtoos-Van Stadens Dunefields (GVD) which is approximately 80km west of the ACD. The minor difference between the two markers regarding location of the phylogeographic boundary is probably due to the dating differences between the two dunefields. The ACD as developed currently is superimposed on the ancient dunefields which formed during the Pleistocene, coinciding with the Cox2-3 spacer sequences divergence which dates back 500,000 - 580,000 years. The GVD formed during the Holocene (6,500 - 4,000 years ago), coinciding with the ITS1 sequences divergence which dates 4,224 - 4,928 years ago. Thus, these phylogeographic boundaries probably appeared without the influence of biogeographic boundaries, but rather due to the lack of suitable habitat in the dunefields, coupled with short dispersal-distances of the spores. Analysis of the ITS1 and Cox2-3 spacer regions in H. spicifera revealed that the species is characterized by uniform genetic structure along the coastline. This reflects the species`s potential for long range expansion as it inhabits both the intertidal and subtidal zones; and this presumably leads to high gene flow among populations. The ITS1 sequences showed minimal genetic variation of one substitution between the gametophyte and tetrasporophyte generations within H. spicifera. This suggests the predominance of asexual reproduction, which reduces gene flow and fixes alleles between generations. ANOSIM and Bray-Curtis cluster analyses showed scale-dependant variation in the abundances of epifauna (mainly amphipod, isopod, mollusc and polychaete species) on G. pristoides. At small local (within site) and large (among sites) scales, there were weak and no structure in epifaunal abundances respectively. However, at larger, biogeographic scales, samples from the same biogeographic region tended to be clustered together. Thus, there was a group containing predominantly south coast samples and a group containing east coast samples mixed with the remaining south coast samples. Such scale-dependant variation in epifaunal abundances is probably due to the effects of factors driving species richness at small local (within site) scales (e.g. wave exposure, seaweed biomass) and at larger, biogeographic scales (e.g. surface sea temperature). Moreover, at very small (individual samples) scales; there was no correlation between epifauna composition and genotype of the seaweed. Seaweed samples characterized by distinct ITS1 or Cox2-3 spacer sequences did not show any significant differences in epifaunal composition. Although the distributional pattern of the epifaunal community observed at large biogeographic scale is not clear, it seems to be associated with the biogeographic regions. However, phylogeographic distribution of Gelidium pristoides is not connected to biogeographic regions. Thus, at larger, biogeographic scales, there is no correlation between phylogeographic distribution of G. pristoides and distribution of the associated fauna

The decrease of biological diversity from low latitudes near the equator towards high latitudes is one of the most fundamental patterns noted in ecology. These trends have been the subject of numerous terrestrial and marine studies and have caused the proposal of several explanatory hypotheses. No single hypothesis has adequately accounted for these observed trends. Furthermore, large-scale patterns are frequently modified by processes acting at meso- to small-scales. It is imperative to understand the interaction of these processes to elucidate the mechanisms controlling the structure of intertidal assemblages. The main aim of this thesis was to test the influence of multiple processes at a range of spatial scales on biogenic engineers. Biogenic engineers, such as macroalgae, have been reported to be strongly influenced by processes such as grazing, biogeography and upwelling and subsequent effects are expected to be linked to their associated assemblages. I used infaunal assemblages associated with macroalgae as a model system to understand the interactive effects of meso-scale upwelling in conjunction with large-scale factors (regional and biogeographic). Three species of macroalgae, Hypnea spicifera, Champia lumbricalis and Bifurcaria brassicaeformis, inhabiting different but overlapping home ranges were used in this study. Smaller scale, physical attributes and the associated fauna of these algal species were compared. The effects of meso- to large-scale physical gradients on marine organisms have been investigated in many instances and resultant gradients in physical variables observed. This study was undertaken to investigate morphological trends exhibited by Hypnea spicifera and Champia lumbricalis to compare possible similarities between coastlines and to determine if inferred meso- to large-scale processes influence physical trends in a similar manner despite the coastlines being affected by different hydrodynamic processes. The macroalgae were located in two distinctly different upwelling regimes, Hypnea spicifera on the east and south coasts and Champia lumbricalis on the west coast. Upwelling (delivery of nutrients and primary production) was not found to be an important factor in determining the physical characteristics of macroalgae but there were clear patterns linked to latitude. Mean percentage cover of Hypnea spicifera was positively correlated with distance along the east and south coasts. Mean mass and mean surface area of H. spicifera were, however, negatively correlated with distance from Port St Johns. Mean frond length showed a negative trend but this relationship was not statistically significant. On the west coast, there was a significant increase in mean frond length from north to south for Champia lumbricalis. Mean surface area and mean mass of C. lumbricalis exhibited non-significant negative trends from north to south while there was a non-significant positive trend for mean percentage cover. Following the “Productivity Hypothesis”, these patterns from north to south along the coastlines of South Africa could have important implications for biodiversity associated with these algae. Many previous studies have focussed on the effects of upwelling on species (i.e. effects of nutrients and temperature) but have failed to separate this effect from large scale effects such as biogeography and latitudinal gradients. I tested the influence meso-scale upwelling, large-scale biogeographic processes and latitudinal gradients with two different species of macroalgae one on the east and south coasts of South Africa and the other on the west coast. Hypnea spicifera inhabits the east and south coasts of South Africa spanning two biogeographic provinces and is generally affected by relatively weak upwelling, whereas Champia lumbricalis inhabits the west coast spanning one biogeographic province which is subjected to intense persistent upwelling year round. Within the east and west coast biogeographic provinces there are, however, regions with both upwelling and non-upwelling. On the east and south coasts, entire assemblages differed significantly among the three Regions (St Lucia, Port Alfred and Knysna, while assemblages between upwelling/non-upwelling areas were not different. Assemblages on the different shores differed significantly from each other. There were no significant effects of region or upwelling for the number of individuals of Crustacea or Polychaeta, while Mollusca showed a significant effect of region. The number of individuals of other taxa showed a significant interaction of region and upwelling. Region had a significant effect on number of species of molluscs and other taxa, while there was no effect for either region or upwelling for the Crustacea or Polychaeta. In general different factors were shown to be important (region and shore) while upwelling was rarely important. On the west coast assemblages associated with C. lumbricalis were not influenced by region or upwelling but there were significant differences between shores. Region, upwelling ad shore did not affect number of individuals. There was a significant interaction of region and upwelling for the number of species of crustaceans, while numbers of species of Mollusca, Polychaeta and other taxa showed no effects. In general, regional factors strongly influenced most organisms, while upwelling played a minor role. Hypotheses about differences between upwelling and non-upwelling regions in terms of species abundances and composition due to the input of cold nutrient rich water were not supported. A link between the effects of larger-scale biogeographic factors and their influence on habitat forming taxa and the resultant effects on associated infauna would be a likely explanation for the patterns observed in this study. Diversity of assemblages is known to be influenced by the structural complexity of a habitat, increasing complexity increases the amount of available niches therefore potentially increasing the number of species found within that habitat. Three species of macroalgae, Hypnea spicifera, Champia lumbricalis and Bifurcaria brassicaeformis were selected on the basis of being structurally similar and having their distributional ranges overlap in order to elucidate the effects of structure and macroalgal species on associated assemblages. Hypnea spicifera is different in terms of surface area and biomass from Champia lumbricalis and Bifurcaria brassicaeformis. Assemblages of species found on the three rhodophytes showed both shore and the species of alga had significant effects on composition. Only numbers of individuals of molluscs were affected by the species of alga. Crustacea and Polychaeta showed an interaction of the species of alga with shore. In the case of this study, it is likely that the three species of macroalgae mitigate biological stressors such as predation and physical stressors such as wave exposure and desiccation. In general, meso-scale upwelling is suggested to be marginalised when considering the structuring of assemblages associated with macroalgae, while large scale biogeography has more of an influence. Within shores, assemblages were also strongly influenced by smaller-scale factors such as differences in the structure and species of alga. This study indicates the importance of investigating patterns across a range of spatial scales to gain a comprehensive understanding of factors influencing intertidal organisms.

The brine shrimp lethality assay was used as a preliminary tool to screen eighteen seaweeds collected from the South African coast. Of the seaweeds tested, the red algae Plocamium corallorhiza and Hypnea rosea, and the green alga Halimeda sp., showed the most potent activity. The chemical investigation of P. corallorhiza resulted in the isolation and structural elucidation of five previously undescribed secondary metabolites, along with three known compounds and four possible artifacts of the extraction process. Standard spectroscopic methods and comparison with known compounds were used to determine the structures of the new metabolites. The new compounds included the linear halogenated monoterpenes 4,8-dibromo-1, 1-dichloro-3,7-dimethyl-2,6-octadiene (99), 4,6-dibromo-l, 1-dichloro-3,7-dimethyl-2,7-octadiene (100), 4,8-dibromo-l, 1,7-trichloro-3,7-dimethyl-2,5-octadiene (101) and 3,4,6,7-tetrachloro-3,7-dimethyl-l-octene (102) and the cyclic monoterpene 5-bromo-5-bromomethyl-I-chlorovinyl-2,4-dichloro-methylcyclohexane (103) while the known compounds were identified as 4-bromo-5-bromomethyl-1chlorovinyl-2,5-dichloro-methylcyclohexane (35), 1,4,8-tribromo-3, 7 -dichloro-3,7-dimethyl-1,5-octadiene (94) and 8-bromo-1,3,4,7-tetrachloro-3,7-dimethyl-1,5-octadiene (96). The four methoxylated compounds (104-107) were presumably formed via a standard substitution reaction between the halogenated monoterpenes 96 and 101 and MeOH, which was used as a component in the extraction solvent. With over 100 000 natural products having been reported, it has become necessary to employ an efficient dereplication strategy to quickly identify known compounds. A simple Gas Chromatography-Mass Spectrometry (GC-MS) method for the efficient physicochemical screening, identification and dereplication of Plocamium metabolites was developed. In this study the crude extracts of P. corallorhiza, P. cornutum and P. maxillosum were screened by GC-MS and the retention times and mass spectral fragmentation patterns of compounds 94, 96, 99 - 107 were used to quickly identify known and new compounds in the crude extracts of P. cornutum and P. maxillosum. This data indicated that compounds 99, 100, 103 were present in both P. corallorhiza and P.cornutum, while compound 102 was found to be present in P. corallorhiza, P. cornutum and P. maxillosum. These studies also indicated that ecotypes and chemotypes are not a significant feature of P. corallorhiza and P. cornutum. Different species of Plocamium (namely: P. corallorhiza, P. cornutum, and P. maxillosum) have very different chemical profiles, and GC may therefore have appreciable taxonomic application in the identification of the different Plocamium spp. which are endemic to South Africa.

The ecophysiology of the red alga Gelidium pristoides (Turner) Kuetzing was investigated in an effort to establish a technique for commercial cultivation. The seaweed is of commercial importance in South Africa where it is harvested from the intertidal zone rocky shores along the coast. It is dried and exported abroad for the extraction of agar. Yields and quality could be improved by cultivation in commercial systems. However, attempts at growing the seaweed in experimental systems have all ended in failure. This study aimed to describe the conditions in which the seaweed grows naturally; and investigate its physiological response to selected physical conditions in the laboratory in order to determine suitable conditions for mariculture. Ecological studies showed that G. pristoides grew above the spring low tide water level. The upper limit of the seaweed’s vertical distribution range, as well as its abundance, was largely dependent on wave exposure. The zone normally inhabited by G. pristoides was dominated by coralline turf in sheltered areas, while the abundance of G. pristoides increased towards more exposed rocky shore sites. The seaweed occurred among species such as Pattelid limpets and barnacles, but was usually the dominant macroalga in this zone, with coralline turf and encrusting algae being the only others. Physical conditions in the part of the intertidal zone inhabited by G. pristoides were highly variable. During low tide temperatures could vary by as much as 10°C within the three hours between tidal inundation of the seaweed population, while salinity varied by up to 9 ppt, and light intensity by as much as 800 μmol m-2 s-1. During these exposure periods the seaweed suffered up to 20% moisture loss. Laboratory experiments on the seaweed’s response to these conditions indicated that it was well adapted to such fluctuations. It had a broad salinity (20 and 40 ppt), and temperature tolerance range (18 to 24°C), with an o ptimum of temperature of 21°C for photosynthesis, while there was no difference in the photosynthetic rate of the alga within the 20 to 40 ppt salinity range. The alga had a low saturating irradiance (ca. 45 – 80 μmol m-2 s-1) equipping it well for photosynthesis in turbulent environments, with high light attenuation, but poorly to unattenuated light conditions. Exposure resulted in an initial increase in photosynthetic rate followed by a gradual decrease thereafter. pH drift experiments showed that low seawater pH, and associated increased carbon dioxide availability, resulted in an increase in photosynthetic rate. This response suggests that the seaweed has a high affinity for carbon dioxide, while the reduction in photosynthetic rate in response to bicarbonate use inhibition indicates that it also has the capacity for bicarbonate use. The high affinity of Gelidium pristoides for carbon dioxide as an inorganic carbon source appears to be the primary reason for the low abundance of the alga on sheltered rocky shore areas, and also explains the failure of the alga to grow in tank or open-water mariculture systems. Exposed rocky shores have experience heavy wave action, and the resultant aeration and mixing of nearshore waters increases the availability of carbon dioxide, which is considered a limiting resource. The absence of such mixing and aeration at sheltered site makes this less suitable habitat for G. pristoides. Periodic exposure also makes high levels of atmospheric carbon dioxide available from which the seaweed benefits. The traditional mariculture systems in which attempts have been made to cultivate the seaweed failed to satisfy either of the above conditions.

Thesis (MTech (Oceanography))--Cape Peninsula University of Technology, 2010 / Coral reefs are in a worldwide state of crisis due to overexploitation. One of the factors contributing to the over exploitation is the marine ornamental industry. This industry utilises a diversity of organisms, comprising approximately 1470 species of fish and more than 300 species of invertebrates. Between April and December 2007, five of the 18 companies who had import permits for South Africa, imported a total of 359 different ornamental species, 252 of these were fish species while the remaining 107 species were invertebrates. During these nine months a total of 32 005 individual organisms were imported into South Africa by the five companies. There were two groups of fish that represented the highest import volumes. These were Amphiprion spp. and Pomacentradae spp., as well as the Gobidae spp. These groups represented 38% and 19% respectively of the total number of fish imported" Aquaculture has the potential to substantially reduce the harvesting pressure on coral reef organisms globally. A small-scale multi-species mariculture system was designed and constructed for benthic egg-laying species of fish (brood stock), corals, and ornamental algae. The system was designed to be low cost and easy to operate. The total capital costs for the system was R15 680.70. In order to estimate the potential yield of an aquaculture facility it is important to know the growth rate of the proposed species under pilot conditions. The use of artificial lighting regimes in aquaculture comes at a financial cost. It is for this reason that it is important to know what lighting scenario yields the highest growth rate of corals. The effects of photoperiod were tested on Sinularia sp. of coral. Two photoperiods were tested, namely: Groups (8:16 h Iight:dark cycle) and Group12 (12:12 h light:dark cycle). No significant difference in weight was found between Group8 and Group12 (p = 0.975). There was however a significant increase (p = 0.002) in Surface-area Pixel Value (SPV) for Groups (1996.73 millipixels pixel-1 day -1) compared to Group12 (983.73 millipixels pixel-1 day -1). The use of a 8:16 h lightdark cycle can thus yield coral of a larger size but not necessarily a higher mass.

Secondary metabolites from natural sources are fast growing as popular drug leads. The structural novelty and favourable biological activity that these compounds display contribute to their popularity as drugs of the future. Examples of such compounds include the potent anticancer drug paclitaxel isolated from the bark of a yew tree as well as the more commonly known analgesic aspirin which stems from the bark of the willow tree. The biological activities exhibited by these secondary metabolites are vast and range from antimicrobial to anticancer activity to mention but a few. As a result, the isolation of novel compounds from natural sources is on the rise. The South African seaboard is home to a wealth of various marine algal species which produce fascinating secondary metabolites. For example, Portierria hornemanii was shown to produce halomon, a halogenated monoterpene which has displayed promising cytotoxic activity. This study thus focused primarily on pursuing novel compounds from three endemic South African marine algal species which have never been analysed previously from a chemical perspective. These are Plocamium rigidum (Bory de Saint-Vincent), Laurencia natalensis (Kylin) and Delisea flaccida (Suhr) Papenfuss. Four known compounds and one new halogenated monoterpene, (2E,5E,7Z)-8-chloro- 7-(dichloromethyl)-4-hydroxy-3-methylocta-2,5,7-trienal, were isolated from Plocamium rigidum. The breast cancer (MCF-7 cell line) inhibitory activity for these compounds was assessed and it was observed that an increase in the lipophilic nature of the compounds produced more favourable IC50 values. A pre-cursor to bromofucin type compounds, cis-laurencenyne, was isolated from Laurencia natalensis, as well as a new acetoxy chamigrane type compound, 4-bromo- 3,10-dichloro-7-hydroxy-3,7,11,11-tetramethylspiro [6.6] undec-1-yl acetate. Delisea flaccida was seen to contain two known bromofuranone type compounds isolated as an isomeric mixture, 1-[(5Z)-4-bromo-5-(bromomethylidene)-2-oxo-2,5- dihydrofuran-3-yl] butyl acetate and 1-[(5E)-4-bromo-5-(bromomethylidene)-2- oxo-2,5-dihydrofuran-3-yl]butyl acetate. These compounds are famous for their ability to inhibit bacterial biofilm production and they have been isolated before from an Australian Delisea spp / Adobe Acrobat 9.53 Paper Capture Plug-in