M. Tech (Biosciences, Faculty of Applied and Computer Sciences), Vaal University of Technology. / The mass production of algae for commercial purposes has predominately been carried out in open ponds systems. However, open ponds systems have a number of disadvantages such as poor light utilization, requirement for large areas of land and high risks of contamination. On the other hand, photobioreactors have attracted much interest because they allow a better control of the cultivation conditions than open systems. With photobioreactors, higher biomass productivities are obtained and contamination can be easily prevented. Photobioreactors can also be engineered to manipulate the light and dark photosynthetic reactions thus enhancing biomass productivity. The main objective of this study was to construct a novel tubular photobioreactor which had the ability to expose the cultured alga to light and dark phases with the aim of optimizing the algal biomass production. A novel tubular photobioreactor with the ability to manipulate the cultured alga’s light and dark photosynthetic reactions was constructed in this study. The alga Spirulina platensis was chosen as the test organism in this novel tubular photobioreactor due to a number of reasons such as its globally socioeconomic importance, its tolerance of higher pH and temperature values which makes it almost impossible to contaminate. The cultivation process of Spirulina in the photobioreactor was investigated through alternating light and dark cycles in an attempt to increase the photosynthetic efficiency of the culture. The effect of different light intensities on the growth of Spirulina in the novel tubular photobioreactor was investigated and it was found that the best light condition that favored higher biomass formation was at 600 μ mol m-2 s-1. Five different light/ dark ratios were evaluated at a light intensity of 600 μ mol m-2 s-1 during a batch mode of operation of the novel tubular photobioreactor. The light/ dark ratio of 1:0.25 was found to be the best ratio because it gave the highest biomass in the shortest period of time when compared to the other ratios used. These results seem to suggest that longer light cycle relative to dark cycle results in higher biomass production. The ratio of 1:0.25 was then used to operate the novel tubular photobioreactor in a continuous mode. A maximum biomass productivity of 25 g/m2/day was achieved which corresponded to a net photosynthetic efficiency of 5.7 %. This result was found to be higher than what most photobioreactors could achieve but it was 2.8 g/m2/day lower than the highest ever reported productivity in a photobioreactor when Spirulina is cultivated. The 2.8 g/m2/day lower was attributed to the different materials used in the construction of these two photobioreactors. The photobioreactor which achieved 27.8 g/m2/day was made up of a clear glass whereas the novel tubular photobioreactor was made up of a PVC tubing. PVC tubes tend to change from clear to a milky colour after a certain period when it is used at higher temperature and pH values hence blocks a certain amount of light. Therefore the main recommendation in this study is to use a PVC tubing with a longer life span when used at a higher temperature and pH values.

A thesis submitted to the Faculty of Engineering and the Built Environment, University of the Witwatersrand, Johannesburg, Republic of South Africa, in fulfillment of the requirements for the degree of Doctor of Philosophy in Engineering. Johannesburg, 2016 / The process of photosynthesis is of great importance as it is the reaction of carbon dioxide (CO2) and water with the help of light, ’free’ energy from the sun, to form useful carbohydrates and oxygen. Photosynthesis is therefore useful both in carbon dioxide mitigation and growing bio-feedstocks towards making biofuel. This thesis aims to address two areas for analysing the photosynthesis process: 1. Looking at the physical limits of the growth; and 2. Improving the production rate of some aquatic plants, such as duckweed and microalgae. To address the first aim, the fundamental concepts of thermodynamics were used to analyse the photosynthetic process. It was found that the theoretical minimum number of moles of photons (NP) required (9–17) is less than the values reported by other researchers, suggesting that the photosynthesis process is highly irreversible and inefficient (operating at 35% efficiency or less). This is because the number of moles of photons will increase with greater process irreversibility (when the entropy generated is greater than zero). If the photosynthesis process is indeed that irreversible then the removal of heat (the heat not used by other cellular processes) by the plant becomes a major problem. It is suggested that transpiration, and other cellular processes, are the processes by which that is done, and it is shown that the water needs of the plant for transpiration would dwarf those needed for photosynthesis. Knowing the fundamental limits to growth could also be of use because if an organism was growing at a rate close to this value there would be no advantage to try to do genetic modification to improve its rate. Following the ideas presented above a spectrophotometer was used not only to obtain the absorption spectrum of algae, but it was also used to grow small samples at specific light wavelengths. The algae species researched was Desmodesmus spp., which, for example, is used to remediate waste water or as a source of feedstock for biofuel production. It also tolerates high CO2 concentrations. This simple experimental method demonstrated that a specific light wavelength (in particular the Secomam Prim spectrophotometer) 440 nm was preferred for the algae growth. It was recommended that this specific light wavelength would be best for growth. It might also be useful to know this fact particularly when designing photobioreactors, as this could reduce the amount of heat released into the surroundings and thus make the process more energy efficient. Interestingly, the wavelength for maximum growth corresponded to one of the peaks in the absorption spectra but there was no increase in growth rate corresponding to any of the other peaks. To address the second aim, the author determined how well predictions on improving the growth of algae (Desmodesmus spp. for example), based on a theoretical model, would work when tested experimentally. What the researcher found was that the method improved algae production, using the same set of equipment. The production was improved by a factor of 1.28 and 1.26 (at product concentrations 1000 mg/L and 600 mg/L respectively) when retaining 40% of the algae suspension. The method may be particularly useful when large amounts of biomass are required as there is no extra cost of purchasing additional equipment. The same model was applied to a growth profile of duckweed (Spirodela polyrhiza 8483, which is convertible into biofuel or a source of food), and the author showed that the model could work if the duckweed was provided with an added carbon source. In order to find an economical and reliable alternative to bridge the scale gap between laboratory and industrial production, the author checked if duckweed species (Spirodela polyrhiza 8483, Spirodela polyrhiza 9509, Lemna gibba 8428, Lemna minor DWC 112, Wolffia cylindracea 7340 and Wolffia globosa 9527) could be cultivated in media less expensive than the basal laboratory medium (Schenk and Hildebrandt). The author found that duckweed can be cultivated more efficiently, and in a more cost-effective manner, in the alternative media types, while maintaining growth rates, RGR 0.09 day-1, and starch contents, 5– 17%(w/w), comparable with that obtained with the conventional laboratory media. Thus, by looking at the photosynthesis process thermodynamically and experimentally, it is shown to be possible to improve the process by using concepts presented in this thesis. / MT2017

A the'.is submitted to the Faculty of Science, University of the Witwatersrand, Johannesburg, in the fulfillment of the requirements of the Degree of Magister Scientae. June 1992 / Aspects of the brown alga Pict~'ppt;eris lQ..n..qifolia at. Palm Beach, Natal, were investigated wi\'-;.hspecial reference to the effect of wave expozu~e~ A description of the species is giv~n and the nomencluture discussed. D~strib1i.ltion of n. ).or~gifolia was det~rmimad from literature and herbarium speof.mens . various r(\~:~thodosf measuring wave exposure were assessed and review~~d. At'!;:.emptswere made to quantify sand abrasion and deposition in tihe scudy si.tes. Morphological and structural aspect;s of the cha LLus were found to vary considerably with season and wave exposure" Light and scanning electron microscopy studie.s indicated the relatjvely complex cell organisation of this brown alga. The prolific regeneration from various thallus sections of .Q. longifolia indicate that regenerat.ion is an important survival strate 'JY following damage. Sexual reproduction and tetrasporogenesis were investigated. Sex.ual reproduction was not prolific in the field, however, tetraspores were abundant and are believed to be particul~rly importcmt in high wave exposure areas. 11. _j.Q.ngi;f'clige,x_periences a range of environmental conditions and responds to these with morphological, structural and reproductive &daptations.

Most ecologists work at scales where complexity is greatest ( i.e. local ), and it is not surprising, therefore, that we tend to be captivated by the description and explanation of local variation whilst being pessimistic about the existence of broader patterns. Using a character ( kelp morphology ) known for its local and unaccounted variation, the morphology of the canopy - forming algae Ecklonia radiata ( Phaeophyta ) was quantified across > 5000 km of temperate Australian coastline, ( i ) between different configurations of algal stand ( i.e. monospecific vs mixed - species stands ) and ( ii ) across multiple spatial scales. A key result was that despite variation at local scales ( km ), differences between stands became increasingly clear at broad scales ( 1000 ' s km ), which supports the idea that large - scale patterns can emerge from apparent stochasticity at small scales. Within each stand, regional scale differences in morphological characters were evident ( i.e. Western Australia = South Australia ≠ Eastern Australia ). These characters correlated with geographic and environmental variables to indicate that the majority of morphological variation across temperate Australia was accounted for by longitude, wave exposure, water temperature and plant density. Morphological differences associated with environmental factors may reflect a plastic response to the local environment, or alternatively may reflect genetically fixed traits ( i.e. ecotypes ). An independent test of morphological variation associated with wave exposure environments, using a reciprocal transplant experiment, revealed that morphological plasticity was the mechanism enabling E. radiata to adopt different morphologies between exposure environments. The presence of kelp canopies has strong spatial relationships with organisms growing underneath them, and variation in the morphology of these canopies may facilitate distinct assemblages within the understorey habitat. Variation in the morphology of E. radiata was found to be associated with the structure of understorey assemblages, over broad spatial scales. This canopy - understorey association revealed two ' types ' of kelp forest ; one characteristic of Western and Southern Australia and the other of Eastern Australia. Patterns of canopy - benthos association have mostly been done on horizontal surfaces and experimental tests showed that such patterns on horizontal surfaces were not representative of vertical surfaces, which enables us to recognize the conditions for which we can reliably anticipate the structure of benthic organisms, thereby improving the predictive power of models that account for widespread patterns in subtidal heterogeneity. In conclusion, this thesis suggests that there are fundamental differences between the ecology of kelp forests at local scales ( i.e. between types of stand ) and at regional scales ( i.e. between the south and east coast of temperate Australia ), reflecting differences in kelp morphology that may be caused by environmental conditions ( e.g. exposure ) and may influence associated taxa ( e.g. understorey ). Consideration of such local - scale variation ( specificity ) when testing for the existence of broad - scale phenomena ( generality ) not only strengthens our understanding of the ecology of subtidal forests, but will also improve the predictive power of further research in this system. / Thesis (Ph.D.)--School of Earth and Environmental Sciences, 2005.

abstract: An issue with the utilization of swimming pools is that pumps are operated an excessive number of hours to keep the pool free of debris and algae. Case in point, according to the pool industry, a pump should operate one hour for every ten degrees of ambient temperature. A dynamic model and a control strategy have been developed using Matlab/Simulink that uses environmental conditions together with chemicals that hinder or aid algae growth in order to determine algae population. This model suggests ways to function the pump on shorter time intervals to reduce energy consumption, while simultaneously maintaining algae populations at acceptable levels. Other factors included in the model are pool thermal dynamics and pool pump/filter performance characteristics, since they also have an effect algae growth. This thesis presents the first step for an alternative way of operating a swimming pool by minimizing operating costs while eliminating algae. / Dissertation/Thesis / algae simulink model / M.S.Tech Technology 2012

The application of high rate algal ponds (HRAP) in the treatment of brewery effluent that met the South African Department of Water Affairs and Forestry's (DWAF) general limits for discharge into a natural water resource of 1998 were tested during a lO-month baseline phase, followed by an 11-month optimization phase. The objective of the baseline phase was to monitor the seasonal performance of HRAPs. The hydraulic retention time (HRT) fluctuated between 11.16 d and 12.00 d in HRAPs. The chemical oxygen demand (COD) increased from 130.12 ± 6.94 mg/L (post-AD), to 171.21 ± 7.99 mg/L (post-HRAP) . The presence of algal cells and evaporation contributed towards an increase in post-HRAP COD. The ammonia (NH₄-N) concentration decreased from 46.59 ± 2.47 mg/L (post-AD), to 1.08 ± 0.12 mg/L (post-HRAP). The nitrite (NO₂- N) concentration remained below 1.00 mg/L in post-pilot plant AD, post-PFP and post-HRAP effluent. The phosphate (PO₄-P) concentration decreased from 29.81 ± 1.39 mg/L (post-AD) to 17.30 ± 1.16 mg/L PO₄-P. The objective of the optimization phase was to manipulate the HRT to achieve the maximum treatment rate that met the DWAF general limits for discharge into a natural water resource of 1998. Nitrogen (as NH₄-N, NO₃-N, NO₂-N) removal efficiency was used as an indicator of nutrient removal success. HRT was influenced by season. The optimal HRT for autumn was 4.30 d at a temperature of 20.53ºC in HRAP A2 (heated) and 18.96ºC in HRAP B2 (ambient). The optimal HRT for summer was 2.74 d at 29.90ºC in HRAP A2 (heated) and 26.36ºC in HRAP B2 (ambient). The COD decreased from 152.33 ± 4.85 mg/L (post-AD) to 95 .00 ± 3.75 mg/L (post-HRAP A2), and to 100.82 ± 5.93 mg/L (post-HRAP B2). The incoming NH₄-N concentration decreased from 42.53 ± 1.38 mg/ L (post-AD), to 1.70 ± 0.81 mg/ L (post-HRAP) . The nitrate (NO₃-N) concentration post-HRAP was 12 - 14 mg/L. The main methods for NH₄-N removal were probably NH₄-N volatilization through algal uptake. HRAPs were able to lower nitrogen and phosphorous concentrations to within the DWAF limits under normal operating conditions. It is recommended that HRAP treated brewery wastewater be used for irrigation after salt removal, or alternatively, for groundwater recharge . Regulatory exemptions would be required for higher than permitted COD and EC concentrations to enable these actions.

Breast cancer, the most common malignancy diagnosed in women, is one of the leading causes of death in women worldwide. In South Africa only 32% of women diagnosed with advanced breast cancer survive more than five years. The search for new chemotherapeutic agents capable of effectively treating breast cancer is therefore essential. Recent evidence supporting the cancer stem cell theory of cancer development for breast cancer challenges the current theories of cancer development and hence treatment. Cancer stem cells are a small subpopulation of tumour cells that possess properties of both cancer cells and stem cells and are believed to be the tumour-initiating population of many cancers. Cancer stem cells are inherently resistant to many chemotherapeutic agents and in this way have been associated with repopulation of tumours after chemotherapy. This phenomenon is proposed as a possible mechanism for cancer relapse after treatment. Cancer stem cells have also been implicated in metastasis, the major cause of mortality in cancer patients. Therefore, any treatment that is capable of targeting and removing breast cancer stem cells may have the theoretical potential to effectively treat breast cancer. However, there are currently no such treatments available for clinical use. We were provided access to a library of novel indigenous small molecules isolated from red and brown algae found off the Eastern Cape of South Africa. The aim of this project was to analyse the anti-cancer and anti-cancer stem cell properties of the compounds in this library and to identify „hit‟ compounds which could form the basis for future development into new anti-cancer drugs. Ten novel compounds of algal origin were tested for cytotoxicity, by determining their ability to inhibit the growth of MCF12A breast epithelial cells and MCF7 breast cancer cells using the colorimetric MTT [(3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] cell proliferation assay. All but one of the compounds tested exhibited cytotoxicity towards the MCF7 cancer cell line, with IC50 values (the concentration of the compound that leads to a 50% inhibition in cell growth) of between 3 μM and 90 μM. The chemotherapeutic drug paclitaxel was used as a positive control. Four of the compounds (RUMB-001, RUMB-002, RUMB-007 and RUMB-010/saragaquinoic acid) were significantly more toxic to the MCF7 cancer cell line, than the „normal‟ MCF12A breast cells and were selected as priority compounds for further analyses. In addition, two other compounds were selected as priority compounds, one highly cytotoxic towards both MCF12A and MCF7 cell lines (RUMB-015) and one which was non toxic to either cell line (RUMB-017/018). Preliminary studies into the mechanism of cytotoxicity using Western blot analysis for poly (ADP-ribose) polymerase (PARP) cleavage and Hoechst 33342 immunostaining in MCF-7 cells were largely unsuccessful. The Hoechst 33342 immunostaining assay did provide tentative evidence that selected priority compounds were capable of inducing apoptosis, although these assays will need to be repeated using a less subjective assay to confirm the results. The priority compounds were subsequently investigated for their cytotoxic effect on the cancer stem cell-enriched side population in MCF7 cells. The ability of the priority compounds to selectively target the cancer stem cell containing side population was assessed using two complementary flow cytometry-based techniques – namely the Hoechst 33342-exclusion assay, and fluorescent immunostaining for the expression of the putative cancer stem cell marker, ABCG2+. The ABCG2+ staining assay was a novel technique developed during the course of this study. It remains to be fully validated, but it may provide a new and reliable way to identify and analyse cancer stem cell containing side population cells. The MCF7 cells were treated with the compounds and the proportion of putative cancer stem cells compared with the size of the population in untreated cells was assessed. Three compounds (RUMB-010, RUMB-015 and RUMB-017/018) capable of reducing the proportion of side population cells within the MCF7 cell line were identified. Taking these data together, we identified two potential „hit‟ compounds which should be prioritised for future research. These are compounds RUMB-010/sargaquinoic acid and RUMB-017/018. RUMB-010 is of interest as it was shown to target the putative cancer stem cell population, in addition to the bulk MCF7 tumour line, but was relatively less toxic to the „normal‟ MCF12A cell line. RUMB-017/018 is of interest due to the ability to selectively target the cancer stem cell enriched side population, while having little effect on the normal (MCF12A) or bulk tumour (MCF7) cell lines tested. These compounds will be important as „hit‟ compounds for drug development and as tool compounds to study cancer and cancer stem cell biology.

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.

Kepone®, the registered trade name for decachlorooctahydro-1,3, 4-metheno-2H-cyclobuta cd pentalen-2-one, inhibited growth and reduced rates of photosynthesis and respiration of Chlorella sorokiniana Shihira and Kraus and Chlorocbccum hypnosporum Starr. The insecticide reduced rates of respiration more than photosynthesis. Uptake of ¹⁴C-Kepone by the algae was proportional to cellular concentration. A net efflux of Kepone was exhibited by Chlorella cells after 30 minutes of incubation, whereas, equilibrium occurred within 15 minutes of incubation for Chlorococcum. Desorption of Kepone was evident when both algal species were removed from Kepone treated solutions and incubated in untreated nutrient solutions. / Master of Science