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.

No description available.

This study was conducted at the installation of the Rural Academic Unit-Tiahuanaco of the BCU, located in the community of Achaca, third municipal section of Ingavi province, department of La Paz. It is 57 km from the La Paz-Desaguadero international highway, at 68 degrees 42 minutes 28 seconds latitude South by 16 degrees 35 minutes 41 seconds longitude West, at an altitude of 3856 meters above sea level. The study went from October 2000 to September 2001. The study consisted of determining the concentrations of the biochemical components in llama seminal plasma at three ages. Components studied were glucose, inorganic phosphate, creatinine, total protein, albumin, globulins, cholesterol, calcium, potassium, sodium, and magnesium. Twelve male llamas of 3, 4, and 5 years were selected and acquired from the Choquecota area of the Carangas province, department of Oruro. Four animals were chosen at each age and were subjected to a training period of semen collection during 2 months, using the artificial hindquarters designed for this effect. The 6 that best responded to the training were used for the investigation. Eight collections were obtained from each animal over the course of the study, and they were used for laboratory analysis. The results were analyzed using a hierarchical factorial design that involved a mixed analysis (nested and crossed) of the factors of age and collections. (The averages of two collections corresponding to each week were analyzed.) Each weekly collection average per age was an experimental unit. Four experimental units were obtained for each age, and the analysis of the data was done with the SAS statistics package version 6.12. From the analyses done the following results were obtained: the concentrations of glucose (6.246 [plus or minus] 0.716 mg/dl), creatinine (3.459 [plus or minus] 1.27 mg/dl), cholesterol (67.28 [plus or minus] 18.21 mg/dl), potassium (8.249 [plus or minus] 1.78 mEq/L), and sodium (123.187 [plus or minus] 18.39 mEq/L) did not show significant differences between ages or collections (p>0.05). The concentrations of calcium (12.138 [plus or minus] 3.64 mg/dl) and magnesium (1.943 [plus or minus] 0.52 mEq/L) showed significant (p<0.05) differences in age only and not in collections. Globulins (1.574 [plus or minus] 0.51 g/dl) showed differences between collections (p<0.05) but not between ages. Total protein (3.732 [plus or minus] 0.45 g/dl), albumin (2.158 [plus or minus] 0.46 g/dl), and inorganic phosphate (9.42 [plus or minus] 2.42 mg/dl) showed differences both between ages and between collections (p<0.05).

The metabolism of white-rot fungi has many proposed biotechnological applications. Their unique capability to depolymerize and catabolize lignin, the most recalcitrant component of lignocellulosic biomass, could be instrumental to the sustainable production of fuels, chemical, and materials from waste biomass feedstocks. The non-specific, oxidative nature of this lignin-degrading metabolism of white-rot fungi renders them capable of degrading a wide range of complex refractory organic compounds beyond lignin, including emerging micropollutants such as pharmaceuticals and pesticides which current wastewater treatment processes were not designed to remove. However, harnessing these metabolic capabilities into engineered bioprocesses has proven to be challenging. Common bioreactor design strategies were developed for traditionally-used unicellular bacteria and yeasts and are not necessarily appropriate for the more complex, filamentous white-rot fungi. Due to a lack of specific engineering strategies and other knowledge gaps, the realization of white-rot fungal bioprocesses has been hampered by low process efficiencies and operational challenges. This dissertation aims to expand the engineering toolbox for harnessing the metabolism of white-rot fungi in bioprocesses. Specifically, it proposes the addition of Fenton chemistry as an avenue to unlock the biotechnological potential of white-rot fungi. The production of hydroxyl radicals through the Fenton reaction is generally understood to be part of the lignin-degrading machinery of white-rot fungi and the addition of Fenton chemistry has been shown to synergistically enhance lignin degradation by white-rot fungi. Overall, the research presented here aims to demonstrate that incorporating Fenton chemistry into white-rot fungal bioprocesses not only synergistically increases lignin degradation efficiency, but also offers a potential solution for the operational challenges that have prevented the implementation of white-rot fungal bioprocesses. This dissertation was guided by five objectives aimed at illustrating the utility of coupling Fenton chemistry and white-rot fungi in engineered bioprocesses. The first objective was to demonstrate, optimize, and uncover the underlying mechanisms driving the synergistic degradation of lignin by white-rot fungi and Fenton chemistry. Through this assessment, it was found that lignin degradation increased synergistically from 58.8% to 80.2% in the presence of Fenton chemistry at the optimum concentration. This work also showed that Fe(II)/Fe(III) cycling and the induction of auxiliary ligninolytic pathways mediate this synergistic interaction. The second objective was to elucidate how Fenton chemistry influences the regulating mechanisms of ligninolytic activity in white-rot fungi, specifically C:N ratio. This showed that C:N ratio significantly influences lignin degradation in the absence of Fenton, but that this effect is blunted in the presence of Fenton. The third objective was to investigate how Fenton chemistry modulates the relationship between the concentration of fungal biomass and the extent of lignin. In the absence of Fenton, fungal biomass concentration was strongly correlated to the extent of lignin degradation. While this was also the case in the presence of Fenton chemistry at very low fungal biomass concentrations, this relationship became uncoupled at sufficiently high fungal biomass concentrations. The fourth objective was to evaluate Fenton chemistry as a selective disinfectant to allow for the persistence or enrichment of white-rot fungi in non-sterile settings. The model competitor E. coli became completely inactivated within hours at the optimal concentration of Fenton reagents, whereas the white-rot fungus P. chrysosporium survived and grew. Lastly, the fifth objective was to demonstrate the long-term performance of a continuously-operated bioreactor which integrated Fenton chemistry and white-rot fungal metabolism. A rotating biological contactor (RBC) combined with a rotating cathode electro-Fenton was constructed and a kinetic model based on batch tests was successfully developed and validated. The reactors were operated for over 100 days and reached stable lignin degradation performance at ~55%. Analysis of the microbial ecology of these reactors showed the persistence of the inoculated P. chrysosporium within the biofilms, as well as the enrichment for other lignin-degrading fungi and bacteria with aromatic catabolism and iron-reduction capabilities. Overall, this research provides insight into the potential and practical implications of integrating Fenton chemistry with white-rot fungi in bioprocesses. The lignin-degrading metabolism of white-rot fungi has long been of interest for biotechnological purposes, but attempts to operationalize them have thus far been unsuccessful at scale. In order to consider scaling white-rot fungi to full-scale operations such as wastewater treatment plants, a better understanding and tighter controls on the growth, ligninolytic activity, and ecological interactions of white-rot fungi are needed. This work proposes Fenton chemistry as a synergetic actor, selective promoter and regulator of white-rot fungal biomass and their production of lignin degrading enzymes.

Utilizou-se um meio de cultivo a base de farinha de mandioca, suplementado com nutrientes, em fermentador agitado (700 rpm) e aerado (10 litros de ar/min), com volume de reação de 10 litros praticamente constante, fração de inóculo de 10% em volume, ph 4,0 e temperatura de 35ºC. Foram realizados ensaios com concentração total de açúcares de 20 g/l e 40 g/l, tanto descontínuos como descontínuos alimentados. Nestes variou-se a vazão mássica de alimentação (fs), o instante de início de alimentação e a condição do xarope de farinha (previamente hidrolisado ou não). Repetições dos ensaios descontínuos indicaram variabilidade de resultados elevada. Não se observou expressivas mudanças no crescimento microbiano, a não ser pelo aumento na velocidade específica nos ensaios descontínuos alimentados a 20 g/l. A síntese de glicoamilase foi sensivelmente aumentada nos ensaios descontínuos alimentados a 20 g/l (produtividade dobrada). A 40 g/l, obteve-se produtividade 26% superior. Os melhores resultados foram obtidos com fs=17,1 gart/h a so=20 g/l e fs=32,2 gart/h a 40 g/l, e obteve-se o pior no ensaio em que se alimentou desde o início de cultivo. A so=20 g/l a repressão se apresenta como principal mecanismo de controle de síntese de glicoamilase, não ocorrendo a mesma a 40 g/l, ensaios nos quais a indução tornou-se muito relevante. / In order to study different processes and the influence of control mechanism on glucoamylase synthesis, several batch and fed-batch runs were made with Aspergillus awamori NRRL 3112. A medium containing cassava flour and nutrients were used in a 10 liters stirred and aerated tank, at pH 4,0 and temperature 35 °C. The batch and fed-batch runs used 20 and 40 g of total reducing sugars (TRS) per liter. In the fed-batch runs, the carbon source feed rate (fs), the feeding start time, and whether the syrup were pre-hydrolyzed or not were varied. Repeated batch runs showed significant variability. Notable changes in cell growth were not observed, unless by the increase of the specific growth rate in the 20 gTRS/l fed-batch runs. The enzyme productivity doubled in the lower sugar concentration fed-batch runs, but increased just 26% in the runs with 40g/l of TRS. The best results were achieved at 20g/l with carbon source feed rate=17,1 gTRS/h and fs=32,2 gTRS/h at 40g/l. The worst noted when the feeding started at the beginning of the run. At 20 gTRS/l, repression showed as the main mechanism control in order to synthesize glucoamylase. On the other hand induction became the relevant factor when 40gTRS/l were offered to microorganism.

In recent years, several industrial yeasts, owing to their robust growth and certain other characteristics, have been developed as recombinant host systems for commercial production of heterologous proteins. One such yeast Pichia pastoris has proven to be an excellent host for production of secreted and intracellular proteins (Cereghino and Cregg. 2000). The increasing popularity of this particular expression system can be attributed to several factors, most importantly: (1) the simplicity of techniques needed for the molecular genetic manipulation of Pichia pastoris and their similarity to those of Saccharomyces cerevisiae, one of the most well-characterized experimental systems in modern biology;(2) the ability of Pichia pastoris to produce foreign proteins at high levels, either intracellularly or extracellularly; and (3) the capability of performing many eukaryotic post-translational modifications, such as glycosylation, disulfide bond formation and proteolytic processing. The expression level for a given recombinant protein produced by Pichia pastoris seems to be determined largely by its inherent properties such as amino acid sequence, the tertiary structure and the site for expression (Sreekrishna et al.,1997). The attempts on increasing the protein expression levels by far are focused on genetic manipulations to enhance the gene expression and protein stability. Although this is crucial, there is ample scope to improve the productivity of Pichia pastoris fermentations by undertaking a systematic program of optimizing the entire fermentation process. This work aims at undertaking such a program by focusing on strategy to identify and to characterize trends in the behavior of the system. It can be expected that by addressing the process as a whole, rather than narrowly focusing on the protein expression alone, the methodology proposed here can simplify process scale-up and can be applied to several products made by the same host. Pichia pastoris is methylotrophic yeast. In the Pichia pastoris fermentation, the limiting carbon source is glycerol, method or mixture of both. It can grow on methanol as a sole carbon and energy source. It possesses a highly inducible methanol utilization pathway. The first step in the metabolism of methanol is the oxidation of methanol to formaldehyde using molecular oxygen by alcohol oxidase (AOX). AOX, the first enzyme of the pathway, accounts for up to 35% of the total protein in cells grown on limited amounts of methanol. The enzymes undetectable in cells grown on glucose, ethanol or glycerol. There are two genes in Pichia pastoris that code for AOX: AOXI. The AOXI gene product accounts for the majority of alcohol oxidase activity in the cell. This highly inducible and stringently regulated AOXI promoter has been used to construct expression vectors for the production of heterologous proteins in Pichia pastoris. Although some foreign proteins have expressed well in shake-flask cultures, expression levels are typically low compared to fomenter cultures. There are several key aspects of Pichia pastoris fermentations: 1. Fed-batch operation – Controlled addition of glycerol, methanol or mixture thereof. In general, strains are grown initially in a defined medium containing glycerol as its carbon source (growth phase). During this phase, biomass accumulates but heterogonous gene expression is fully repressed. Upon depletion of glycerol, a transition phase is initiated in which additional glycerol is fed to the culture at a growth-limiting rate. Finally, method a mixture of glycerol and methanol is fed to the culture to induce expression (induction phase). The duration of individual substrate feeds, the amount and mode of feeding are critical to optimal fermentation performance. 2.Online measurement and control-One of the most important key parameters in Pichia pastor is expression system is the methanol concentration. Monitoring and controlling this variable are important because high levels of this inductor substrate can be toxic to the cells and low levels of methanol may not be enough to initiate the AOX transcription (Cereghino and Cregg, 2000) This research work aims at investigation the above mentioned aspects by conduction an in depth engineering analysis of the Pichia Pastoris fermentations.

This thesis describes the characterization of ceramide (Cer) biosynthesis by mammalian cells. The possibility that Cer undergo developmental changes was explored using mouse embryonic stem cells versus embryoid bodies by analysis of the Cer subspecies by liquid chromatography, electrospray ionization-tandem mass spectrometry (LC ESI-MS/MS) and of the transcript levels for enzymes involved in Cer biosynthesis by qRT-PCR. Cer of embroid bodies had higher proportions of very-long-chain fatty acids, which correlated with the relative expression of mRNA for the respective Cer synthases (CerS) and fatty acyl-CoA elongases, as well as changes in the fatty acyl-CoA's of the cells. Therefore, it is clear that Cer subspecies change during embryogenesis, possibly for functionally important reasons. One CerS isoform, CerS2, was studied further because it has the broadest tissue distribution and a remarkable fatty acyl-CoA specificity, utilizing longer acyl-chain CoAs (C20-C26) in vitro. The fatty acid chain selectivity was refined by analysis of the Cer from livers from CerS2 null mice, which displayed very little Cer with fatty acyl chains with 24 + 2 carbons. Another interesting structural variation was discovered in studies of cells treated with fumonisin B1 (FB1), which inhibits CerS. Under these conditions, cells in culture and animals accumulate substantial amounts of a novel sphingoid base that was identified as 1-deoxysphinganine. This compound arises from utilization of L-alanine instead of L-serine by serine palmitoyltransferase (SPT) based on the inability of LYB cells, which lack SPT, to make 1-deoxysphinganine. In the absence of FB1, 1-deoxysphinganine is primarily acylated to 1-deoxydihydroceramides. These are an underappreciated category of bioactive sphingoid bases and "ceramides" that might play important roles in cell regulation and disease. In summary, cells contain a wide variety of Cer subspecies that are determined by changes in expression of CerS, enzymes that produce co-substrates (such as fatty acyl-CoAs), and the types of amino acids utilized by SPT, the initial enzyme of de novo sphingolipid biosynthesis. One can envision how these changes might impact membranes structure as well as signaling by this family of highly bioactive compounds.

Utilizou-se um meio de cultivo a base de farinha de mandioca, suplementado com nutrientes, em fermentador agitado (700 rpm) e aerado (10 litros de ar/min), com volume de reação de 10 litros praticamente constante, fração de inóculo de 10% em volume, ph 4,0 e temperatura de 35ºC. Foram realizados ensaios com concentração total de açúcares de 20 g/l e 40 g/l, tanto descontínuos como descontínuos alimentados. Nestes variou-se a vazão mássica de alimentação (fs), o instante de início de alimentação e a condição do xarope de farinha (previamente hidrolisado ou não). Repetições dos ensaios descontínuos indicaram variabilidade de resultados elevada. Não se observou expressivas mudanças no crescimento microbiano, a não ser pelo aumento na velocidade específica nos ensaios descontínuos alimentados a 20 g/l. A síntese de glicoamilase foi sensivelmente aumentada nos ensaios descontínuos alimentados a 20 g/l (produtividade dobrada). A 40 g/l, obteve-se produtividade 26% superior. Os melhores resultados foram obtidos com fs=17,1 gart/h a so=20 g/l e fs=32,2 gart/h a 40 g/l, e obteve-se o pior no ensaio em que se alimentou desde o início de cultivo. A so=20 g/l a repressão se apresenta como principal mecanismo de controle de síntese de glicoamilase, não ocorrendo a mesma a 40 g/l, ensaios nos quais a indução tornou-se muito relevante. / In order to study different processes and the influence of control mechanism on glucoamylase synthesis, several batch and fed-batch runs were made with Aspergillus awamori NRRL 3112. A medium containing cassava flour and nutrients were used in a 10 liters stirred and aerated tank, at pH 4,0 and temperature 35 °C. The batch and fed-batch runs used 20 and 40 g of total reducing sugars (TRS) per liter. In the fed-batch runs, the carbon source feed rate (fs), the feeding start time, and whether the syrup were pre-hydrolyzed or not were varied. Repeated batch runs showed significant variability. Notable changes in cell growth were not observed, unless by the increase of the specific growth rate in the 20 gTRS/l fed-batch runs. The enzyme productivity doubled in the lower sugar concentration fed-batch runs, but increased just 26% in the runs with 40g/l of TRS. The best results were achieved at 20g/l with carbon source feed rate=17,1 gTRS/h and fs=32,2 gTRS/h at 40g/l. The worst noted when the feeding started at the beginning of the run. At 20 gTRS/l, repression showed as the main mechanism control in order to synthesize glucoamylase. On the other hand induction became the relevant factor when 40gTRS/l were offered to microorganism.

Studies into sources of alternative liquid transport fuel energy have identified agro-industrial wastes, which are lignocellulosic in nature, as a potential feedstock for biofuel production against the background of depleting nonrenewable fossil fuels. In South Africa, large quantities of apple and other fruit wastes, called pomace, are generated from fruit and juice industries. Apple pomace is a rich source of cellulose, pectin and hemicellulose, making it a potential target for utilisation as a lignocellulosic feedstock for biofuel and biorefinery chemical production. Lignocellulosic biomass is recalcitrant in nature and therefore its degradation requires the synergistic action of a number of enzymes such as cellulases, hemicellulases, pectinases and ligninases. Commercial enzyme cocktails, containing some of these enzymes, are available and can be used for apple pomace degradation. In this study, the degradation of apple pomace using commercial enzyme cocktails was investigated. The main focus was the optimisation of the release of sugar monomers that could potentially be used for biofuel and biorefinery chemical production. There is no or little information reported in literature on the enzymatic degradation of fruit waste using commercial enzyme mixtures. This study first focused on the characterisation of the substrate (apple pomace) and the commercial enzyme cocktails. Apple pomace was found to contain mainly glucose, galacturonic acid, arabinose, galactose, lignin and low amounts of xylose and fructose. Three commercial enzyme cocktails were initially selected: Biocip Membrane, Viscozyme L (from Aspergillus aculeatus) and Celluclast 1.5L (a Trichoderma reesei ATCC 26921 cellulase preparation). The selection of the enzymes was based on activities declared by the manufacturers, cost and local availability. The enzymes were screened based on their synergistic cooperation in the degradation of apple pomace and the main enzymes present in each cocktail. Viscozyme L and Celluclast 1.5L, in a 50:50 ratio, resulted in the best degree of synergy (1.6) compared to any other combination. The enzyme ratios were determined on Viscozyme L and Celluclast 1.5L based on the protein ratio. Enzyme activity was determined as glucose equivalents using the dinitrosalicylic acid (DNS) method. Sugar monomers were determined using Megazyme assay kits. There is limited information available on the enzymes present in the commercial enzyme cocktails. Therefore, the main enzymes present in Viscozyme L and Celluclast 1.5L were identified using different substrates, each targeted for a specific enzyme and activity. Characterisation of the enzyme mixtures revealed a large number of enzymes required for apple pomace degradation and these included cellulases, pectinases, xylanases, arabinases and mannanases in different proportions. Viscozyme L contained mainly pectinases and hemicellulases, while Celluclast 1.5L displayed largely cellulase and xylanase activity, hence the high degree of synergy reported. The temperature optimum was 50ºC for both enzyme mixtures and pH optima were observed at pH 5.0 and pH 3.0 for Viscozyme L and Celluclast 1.5L, respectively. At 37ºC and pH 5.0, the enzymes retained more that 90% activity after 15 days of incubation, allowing the enzymes to be used together with less energy input. The enzymes were further characterised by determining the effect of various compounds, such as alcohols, sugars, phenolic compounds and metal ions at various concentrations on the activity of the enzymes during apple pomace hydrolysis. Apart from lignin, which had almost no effect on enzyme activity, all the compounds caused inhibition of the enzymes to varying degrees. The most inhibitory compounds were some organic acids and metal ions, as well as cellobiose and xylobiose. Using the best ratio for Viscozyme L and Celluclast 1.5L (50:50) for the hydrolysis of apple pomace, it was observed that synergy was highest at the initial stages of hydrolysis and decreased over time, though the sugar concentration increased. The type of synergy for optimal apple pomace hydrolysis was found to be simultaneous. There was no synergy observed between Viscozyme L and Celluclast 1.5L with ligninases - laccase, lignin peroxidase and manganese peroxidase. Hydrolysing apple pomace with ligninases prior to addition of Viscozyme L and Celluclast 1.5L did not improve degradation of the substrate. Immobilisation of the enzyme mixtures on different supports was performed with the aim of increasing stability and enabling reuse of the enzymes. Immobilisation methods were selected based on the chemical properties of the supports, availability, cost and applicability on heterogeneous and insoluble substrate like apple pomace. These methods included crosslinked enzyme aggregates (CLEAs), immobilisation on various supports such as nylon mesh, nylon beads, sodium alginate beads, chitin and silica gel beads. The immobilisation strategies were unsuccessful, mainly due to the low percentage of immobilisation of the enzyme on the matrix and loss of activity of the immobilised enzyme. Free enzymes were therefore used for the remainder of the study. Hydrolysis conditions for apple pomace degradation were optimised using different temperatures and buffer systems in 1 L volumes mixed with compressed air. Hydrolysis at room temperature, using an unbuffered system, gave a better performance as compared to a buffered system. Reactors operated in batch mode performed better (4.2 g/L (75% yield) glucose and 16.8 g/L (75%) reducing sugar) than fed-batch reactors (3.2 g/L (66%) glucose and 14.6 g/L (72.7% yield) reducing sugar) over 100 h using Viscozyme L and Celluclast 1.5L. Supplementation of β- glucosidase activity in Viscozyme L and Celluclast 1.5L with Novozyme 188 resulted in a doubling of the amount of glucose released. The main products released from apple pomace hydrolysis were galacturonic acid, glucose and arabinose and low amounts of galactose and xylose. These products are potential raw materials for biofuel and biorefinery chemical production. An artificial neural network (ANN) model was successfully developed and used for predicting the optimum conditions for apple pomace hydrolysis using Celluclast 1.5L, Viscozyme L and Novozyme 188. Four main conditions that affect apple pomace hydrolysis were selected, namely temperature, initial pH, enzyme loading and substrate loading, which were taken as inputs. The glucose and reducing sugars released as a result of each treatment and their combinations were taken as outputs for 1–100 h. An ANN with 20, 20 and 6 neurons in the first, second and third hidden layers, respectively, was constructed. The performance and predictive ability of the ANN was good, with a R² of 0.99 and a small mean square error (MSE). New data was successfully predicted and simulated. Optimal hydrolysis conditions predicted by ANN for apple pomace hydrolysis were at 30% substrate (wet w/v) and an enzyme loading of 0.5 mg/g and 0.2 mg/mL of substrate for glucose and reducing sugar, respectively, giving sugar concentrations of 6.5 mg/mL and 28.9 mg/mL for glucose and reducing sugar, respectively. ANN showed that enzyme and substrate loadings were the most important factors for the hydrolysis of apple pomace.

Thesis (M.Tech.: Chemical Engineering)-Dept. of Chemical Engineering, Durban University of Technology, 2005 1 v. (various pagings) / In bubble-less aeration oxygen diffuses through the membrane in a molecular form and dissolves in the liquid. Oxygen is fed through the lumen side of silicone rubber tube. On the outer surface of the membrane there is a boundary layer that is created by oxygen. This then gets transported to the bulk liquid by convective transport created by water circulation through the pump. The driving force of the convective transport is due to concentration difference between the dissolved oxygen in water and oxygen saturation concentration in water at a particular temperature and pressure. The design of a membrane aerated bioreactor needs an understanding of the factors that govern oxygen mass transfer. It is necessary to know the effects of operating conditions and design configurations. Although various methods of bubble-less aeration have been reported, there still exists a lack of knowledge on the immersed membrane systems. This study is aiming at contributing to the development of an immersed membrane bioreactor using silicone rubber tubular membrane as means of providing oxygen. The secondary objective was to investigate the influence that the operating conditions and module configuration have on the system behaviour. From the experimental study, the characteristic dissolved oxygen -time curve show that there is a saturation limit equivalent to the equilibrium dissolved oxygen concentration, after which there is no increase in dissolved oxygen with time. At ambient conditions the equilibrium dissolved oxygen is approximately 8 mg/L. This is when water is in contact with air at one atmospheric pressure. At the same conditions the equilibrium dissolved oxygen concentration when water is in contact with pure oxygen is approximately 40 mg/L. This is why all the experiments were conducted from 2mg/L dissolved oxygen concentration in water, to enable enough time to reach equilibrium so as to determine mass transfer coefficient. The most important parameters that were investigated to characterise the reactor were, oxygen supply pressure, crossflow velocity, temperature and module orientation. Observations from the experimental study indicated that when the system is controlled by pressure, crossflow does not have a significant effect on mass transfer. When the system is controlled by the convective transport from the membrane surface to the bulk liquid, pressure does not have a significant effect on mass transfer. All four effects that were investigated in the study are discussed.