Thesis (M.Tech.:Biotechnology)-Dept. of Biotechnology, Durban University of Technology, 2007 xvi, 185 leaves / Spirulina is a blue-green, multicellular, filamentous cyanobacterium that can grow to sizes of 0.5 millimetres in length. It is an obligate photoautotroph and has a pH growth range from 8.3 to 11.0.The large-scale production of Spirulina biomass depends on many factors, the most important of which are nutrient availability, temperature and light. These factors can influence the growth of Spirulina and the composition of the biomass produced by changes in metabolism. Brine effluent from cooling towers of electricity generating plants may provide an ideal growth medium for Spirulina based on its growth requirements, i.e. high alkalinity and salinity. The aim of this research was to optimise brine effluent from cooling towers by supplementing it with salts, in order to use this optimised effluent in a small open laboratory raceway pond in an attempt to increase the biomass production of Spirulina.

Thesis (D.Tech.: Biotechnology)-Dept. of Biotechnology, Durban University of Technology, 2007 xi, 209 leaves / Protein engineering technologies, such as directed evolution and DNA recombination, are often used to modify enzymes on a genetic level for the creation of useful industrial catalysts. Pre-treatment of paper pulps with xylanases have been shown to decrease the amounts of toxic chlorine dioxide used to bleach pulp. This study was undertaken to improve the thermal and alkaline stabilities of the xylanase from the fungus Thermomyces lanuginosus using ep-PCR and DNA shuffling.

Thesis (M.Tech.: Biotechnology)- Dept. of Biotechnology, Durban University of Technology, 2007 xv, 127 leaves / The human gut microbiota play a major role in host health, and attempts are being made to manipulate the composition of the gut microbiota-increase the composition of bacterial groups, such as lactobacilli and bifidobacteria that are perceived as exerting health promoting properties. These bacteria defined as food supplements (probiotics) beneficially affect the host by improving the intestinal microbial balance, and have been used to change the composition of the colonic microbiota. However, such changes may be transient, and the implantation of exogenous bacteria therefore becomes limited. In contrast, prebiotics are naturally occurring carbohydrates that are classified as non-digestible oligosaccharides present in edible plants. These carbohydrates enter the colon as intact compounds, elicit systemic physiological functions and act as fermentable substrates for colonic microflora-influencing the species composition and metabolic characteristics of intestinal microflora providing important health attributes. Currently, a widely marketed prebiotic, inulin is extracted from plants of the family Asteraceae. There are many unexploited plants that are regularly consumed and that may have a prebiotic effect or can have very high levels of inulin which could make them commercially viable. In this study, we investigated prebiotic compounds, especially inulin from locally growing, non-commercialised leafy plants. The aqueous extracts of 22 plants from the families Asparagaceae, Alliaceae, Asteraceae, Solanaceae, Cucurbitaceae, Amaranthaceae, Acanthaceae, Polygonaceae, Portulaceae, Fabaceae, Chenopodiaceae, Pedaliaceae and Apiaceae from Kwa-Zulu Natal were investigated for a prebiotic effect using a modified batch-culture technique with Lactobacillus bulgaricus, Lactobacillus lactis, Lactobacillus reuteri and Bifidobacterium longum, four common probiotics and the inulin content of the plants was determined using high performance liquid chromatography. Of the 22 plants studied, Solanum nigrum, Amaranthus spinosus, Amaranthus hybridus, Asystasia gangetica, Senna occidentalis, Cerathoteca triloba, Asparagus sprengeri, Tulbaghia violacea, Sonchus oleraceus and Taraxacum officinale exhibited a prebiotic effect. The prebiotic effect of the Taraxacum officinale, Sonchus oleraceus and Asparagus sprengeri extracts on L. lactis and L. reuteri was higher than or equivalent to inulin-a commercial prebiotic. In this study, Sonchus oleraceus exhibited the best prebiotic effect-was the only plant to stimulate all the probiotics including B. longum. Of all the plants analysed, Asparagus sprengeri tuber contained the highest amount of inulin (3.55%).

Thesis (D. Tech.: Biotechnology)-Dept. of Biotechnology, Durban University of Technology, 2006 xii, 274 leaves / In Sub-Saharan Africa there is an urgent need to sustain and improve the quality of its food resources. Poverty eradication features high on the agenda of a number of world health organisations, while the number of underweight children in Africa continues to increase (Pellet, 1996). Providing nutritionally enhanced foods to the poor will help towards achieving this objective. Protein-energy malnutrition has been identified as one of the most important problems facing Africa, with maize as the staple diet (Nkama et al., 1995). However, a combination of several factors limits availability and the nutritional quality of maize. During starvation, energy and protein intakes decrease by 20-30%, with most of the children in Africa having an average protein intake of only 20 g per day (Igbedioh, 1996). Energy availability also affects protein utilization because of interrelationships of protein and energy metabolism (Elwyn, 1993). The diets of inhabitants in developing regions depend mainly on cereals (maize) for both protein and dietary energy which lacks indispensable amino acids, minerals, vitamins and carbohydrates. In light of these growing concerns an attempt was made to devise a scientific strategy to combat the nutritional shortfalls of maize meal. A multidisciplinary and concerted approach was followed within this project aimed at designing an improved thermostable amylase and applying the enzyme to nutritionally enhance maize meal. It was envisaged that the manipulation of maize meal, by the application of enzyme technology will improve the nutritional status of this staple food. The consequences is that an alternate solution for the eradication of an ailing, poverty stricken and malnourished African population is achievable. It is possible that the boundaries defining the limits of life will extend to even greater extremes through the application of novel technologies.

Thesis (M.Tech.: Food Technology)-Dept of Biotechnology, Durban University of Technology, 2007 xvi, 106 leaves / Alicyclobacillus acidoterrestris is a spore-forming spoilage micro-organism found in fruit juices whose spores are not destroyed by typical pasteurisation. Once its spores outgrow and multiply in finished juice products, they produce two volatile taint compounds namely guaiacol and 2,6-dibromophenol. In the food industry margins for errors are small and monitoring of products to avert such errors is crucial. Conventional microbiological monitoring is one such technique for spoilage micro-organisms another being automated systems which can detect taints. Both these categories were evaluated in this study with the electronic nose and gas chromatograph being the specific automated systems being assessed. Sensory evaluation was also assessed as a diagnostic tool in the detection of taints. Isolation and identification of what was thought to be A. acidoterrestris was a laborious and expensive exercise which eventually proved inconclusive. A pure culture was purchased and juices were then inoculated with two levels of A. acidoterrestris spores and incubated. Juices from each level of inoculation were evaluated at different time intervals via the above-mentioned monitoring techniques. Of the three media assessed in the microbiological method, Bacillus acidoterrestris medium (BAM) was found to be the most effective for enumerating A. acidoterrestris followed by K-medium (KM) then Orange Serum Agar (OSA). While BAM was still indicating the presence of A. acidoterrestris KM and OSA were not (counts of <10cfu/g). This illustrated that this micro-organism could be easily overlooked if KM or OSA were being used to enumerate them. Considering that many workers actually do use KM and OSA as their media of choice in enumerating A. acidoterrestris (perhaps because BAM is very tedious to prepare) the cause for concern is a real one. Assessment of the resultant taints via sensory evaluation after inoculation and incubation reveals the inability of many panellists to detect taints at levels (as assessed by GC) far above their documented threshold values. While GC is an extremely useful and powerful tool, the level of expertise and skill required to use such an instrument cannot be overlooked nor can the expense involved. With regard to the electronic nose assessment for the presence of the volatile taint compounds, an important finding was that the electronic nose indicated significant differences between test and control samples when panellists performing sensory evaluation did not. This also correlated to an interval when enumeration on OSA illustrated no A. acidoterrestris after several days of inoculation and incubation and BAM and KM did. Without implying that the electronic nose has no drawbacks, it has proved, in this instance to be a simple and easy piece of equipment to use. It can be used to detect taints produced under simulated spoilage conditions at reduced analysis times, levels of expertise, cost and energy.

Thesis (M.Tech.: Biotechnology)- Dept. of Biotechnology & Food Technology, Durban University of Technology, 2007 xv, 102 leaves / For decades the genus Aspergillus (of fungi) has been classified based on morphological and growth criteria. Members of the Aspergillus section Flavi are economically valuable and methods of differentiating them are thus very important. Several molecular methods have been developed to distinguish these strains. Also, a number of biochemical and genetic studies have been used in order to provide a better means of classification (Lee et al., 2004). Aflatoxins, the most frequently studied mycotoxins, are produced by certain Aspergillus species/strains/isolates of fungi. The aflatoxin biosynthetic pathway studies have led to a number of discoveries. Several structural and regulatory genes (and their enzymes) involved in the biosynthesis of aflatoxins have been discovered and purified (Trail et al., 1995). Aflatoxin production and contamination of agricultural crops are major causes of economic losses in agriculture. Thus, better methods of characterization/differentiation are required for both aflatoxigenic and non-aflatoxigenic isolates. Molecular biology is one of the current tools used to differentiate between these isolates. Polymerase Chain Reaction (PCR)-based randomly amplified polymorphic DNA (RAPD) analysis has been used successfully in the analysis of DNA relatedness of species of fungi, bacteria, plants and animals. Dendograms which evaluate/assess the likeness between different isolates has also been used (Martinez et al., 2001). Restriction fragment length polymorphism (RFLP) analysis has been applied to a number of studies to detect differences between fungi and to establish relationships between them. Therefore, the scope of this study was to investigate RAPD analysis (with dendograms) and detection of RFLPs by hybridization as molecular methods that can distinctly differentiate or characterize the aflatoxigenic and non-aflatoxigenic Aspergillus isolates.

Thesis (M.Tech.: Biotechnology)- Dept. of Biotechnology, Durban University of Technology, 2007 xv, 127 leaves / The human gut microbiota play a major role in host health, and attempts are being made to manipulate the composition of the gut microbiota-increase the composition of bacterial groups, such as lactobacilli and bifidobacteria that are perceived as exerting health promoting properties. These bacteria defined as food supplements (probiotics) beneficially affect the host by improving the intestinal microbial balance, and have been used to change the composition of the colonic microbiota. However, such changes may be transient, and the implantation of exogenous bacteria therefore becomes limited. In contrast, prebiotics are naturally occurring carbohydrates that are classified as non-digestible oligosaccharides present in edible plants. These carbohydrates enter the colon as intact compounds, elicit systemic physiological functions and act as fermentable substrates for colonic microflora-influencing the species composition and metabolic characteristics of intestinal microflora providing important health attributes. Currently, a widely marketed prebiotic, inulin is extracted from plants of the family Asteraceae. There are many unexploited plants that are regularly consumed and that may have a prebiotic effect or can have very high levels of inulin which could make them commercially viable. In this study, we investigated prebiotic compounds, especially inulin from locally growing, non-commercialised leafy plants. The aqueous extracts of 22 plants from the families Asparagaceae, Alliaceae, Asteraceae, Solanaceae, Cucurbitaceae, Amaranthaceae, Acanthaceae, Polygonaceae, Portulaceae, Fabaceae, Chenopodiaceae, Pedaliaceae and Apiaceae from Kwa-Zulu Natal were investigated for a prebiotic effect using a modified batch-culture technique with Lactobacillus bulgaricus, Lactobacillus lactis, Lactobacillus reuteri and Bifidobacterium longum, four common probiotics and the inulin content of the plants was determined using high performance liquid chromatography. Of the 22 plants studied, Solanum nigrum, Amaranthus spinosus, Amaranthus hybridus, Asystasia gangetica, Senna occidentalis, Cerathoteca triloba, Asparagus sprengeri, Tulbaghia violacea, Sonchus oleraceus and Taraxacum officinale exhibited a prebiotic effect. The prebiotic effect of the Taraxacum officinale, Sonchus oleraceus and Asparagus sprengeri extracts on L. lactis and L. reuteri was higher than or equivalent to inulin-a commercial prebiotic. In this study, Sonchus oleraceus exhibited the best prebiotic effect-was the only plant to stimulate all the probiotics including B. longum. Of all the plants analysed, Asparagus sprengeri tuber contained the highest amount of inulin (3.55%).

Thesis (M.Tech.: Biotechnology)-Dept. of Biotechnology, Durban University of Technology, 2007 xxi, 117 leaves / The aromatic compounds Polychlorinated Biphenyls (PCBs) are one of the largest groups of environmental pollutants. The greatest concern is the release of PCBs in the water systems by industrial effluent, accidental spillages or leaks. PCBs are able to bioaccumulate in the fatty tissues of animals, fish and humans. The impact on human health due to PCBs has prompted interest in their degradation. The application of microbial degradation of PCBs can transform many PCB metabolites. There are a wide variety of microorganisms that can degrade PCBs or utilise them as sole carbon sources. This study focused on isolating microrganisms from industrial wastewater capable of aerobic degradation of PCBs. The degradation potential of the selected isolates were investigated by using different analytical techniques viz. ultra violet or visible spectrophotometer (UV/Vis), thin layer chromatography (TLC) and gas chromatography electron capture detector (GC-ECD).

Thesis (D. Tech.: Biotechnology)-Dept. of Biotechnology, Durban University of Technology, 2006 xii, 274 leaves / In Sub-Saharan Africa there is an urgent need to sustain and improve the quality of its food resources. Poverty eradication features high on the agenda of a number of world health organisations, while the number of underweight children in Africa continues to increase (Pellet, 1996). Providing nutritionally enhanced foods to the poor will help towards achieving this objective. Protein-energy malnutrition has been identified as one of the most important problems facing Africa, with maize as the staple diet (Nkama et al., 1995). However, a combination of several factors limits availability and the nutritional quality of maize. During starvation, energy and protein intakes decrease by 20-30%, with most of the children in Africa having an average protein intake of only 20 g per day (Igbedioh, 1996). Energy availability also affects protein utilization because of interrelationships of protein and energy metabolism (Elwyn, 1993). The diets of inhabitants in developing regions depend mainly on cereals (maize) for both protein and dietary energy which lacks indispensable amino acids, minerals, vitamins and carbohydrates. In light of these growing concerns an attempt was made to devise a scientific strategy to combat the nutritional shortfalls of maize meal. A multidisciplinary and concerted approach was followed within this project aimed at designing an improved thermostable amylase and applying the enzyme to nutritionally enhance maize meal. It was envisaged that the manipulation of maize meal, by the application of enzyme technology will improve the nutritional status of this staple food. The consequences is that an alternate solution for the eradication of an ailing, poverty stricken and malnourished African population is achievable. It is possible that the boundaries defining the limits of life will extend to even greater extremes through the application of novel technologies.

Thesis (M.Tech.: Food Technology)-Dept of Biotechnology, Durban University of Technology, 2007 xvi, 106 leaves / Alicyclobacillus acidoterrestris is a spore-forming spoilage micro-organism found in fruit juices whose spores are not destroyed by typical pasteurisation. Once its spores outgrow and multiply in finished juice products, they produce two volatile taint compounds namely guaiacol and 2,6-dibromophenol. In the food industry margins for errors are small and monitoring of products to avert such errors is crucial. Conventional microbiological monitoring is one such technique for spoilage micro-organisms another being automated systems which can detect taints. Both these categories were evaluated in this study with the electronic nose and gas chromatograph being the specific automated systems being assessed. Sensory evaluation was also assessed as a diagnostic tool in the detection of taints. Isolation and identification of what was thought to be A. acidoterrestris was a laborious and expensive exercise which eventually proved inconclusive. A pure culture was purchased and juices were then inoculated with two levels of A. acidoterrestris spores and incubated. Juices from each level of inoculation were evaluated at different time intervals via the above-mentioned monitoring techniques. Of the three media assessed in the microbiological method, Bacillus acidoterrestris medium (BAM) was found to be the most effective for enumerating A. acidoterrestris followed by K-medium (KM) then Orange Serum Agar (OSA). While BAM was still indicating the presence of A. acidoterrestris KM and OSA were not (counts of <10cfu/g). This illustrated that this micro-organism could be easily overlooked if KM or OSA were being used to enumerate them. Considering that many workers actually do use KM and OSA as their media of choice in enumerating A. acidoterrestris (perhaps because BAM is very tedious to prepare) the cause for concern is a real one. Assessment of the resultant taints via sensory evaluation after inoculation and incubation reveals the inability of many panellists to detect taints at levels (as assessed by GC) far above their documented threshold values. While GC is an extremely useful and powerful tool, the level of expertise and skill required to use such an instrument cannot be overlooked nor can the expense involved. With regard to the electronic nose assessment for the presence of the volatile taint compounds, an important finding was that the electronic nose indicated significant differences between test and control samples when panellists performing sensory evaluation did not. This also correlated to an interval when enumeration on OSA illustrated no A. acidoterrestris after several days of inoculation and incubation and BAM and KM did. Without implying that the electronic nose has no drawbacks, it has proved, in this instance to be a simple and easy piece of equipment to use. It can be used to detect taints produced under simulated spoilage conditions at reduced analysis times, levels of expertise, cost and energy.