published_or_final_version / abstract / Civil Engineering / Doctoral / Doctor of Philosophy

The indiscriminate use of fossil fuels has led to global problems of greenhouse gas emissions, environmental degradation and energy security. Developments of alternative and sustainable energy resources have assumed paramount importance over the past decades to curb these challenges. Biohydrogen is emerging as an alternative renewable source of energy and has received considerable attention in recent years due to its social, economic and environmental benefits. It can be generated by dark fermentation on Organic Fraction of Solid Municipal Waste (OFSMW). These OFSMW exist abundantly and poses disposal challenges. This study models and optimizes the production of biohydrogen on a mixture of agro-municipal wastes; it examines a semi-pilot scale production on these substrates and the feasibility of generating bioelectricity from the process effluents and reviews the prospect of enhancing fermentative biohydrogen development using miniaturized parallel bioreactors. The fermentation process of biohydrogen production on agro-municipal wastes was modelled and optimized using a two-stage design. A mixture design was used for determination of optimum proportions of co-substrates of Bean Husk (BH), Corn Stalk (CS) and OFSMW for biohydrogen production. The effects of operational setpoint parameters of substrate concentration, pH, temperature and Hydraulic Retention Time (HRT) on hydrogen response using the mixed substrates were modelled and optimized using box-behnken design. The optimized mixtures were in the ratio of OFSMW: BH: CS = 30:0:0 and OFSMW: BH: CS = 15:15:0 with yields of 56.47 ml H2/g TVS and 41.16 ml H2/g TVS respectively. Optimization on physico-chemical parameters using the improved substrate suggested optimal setpoints of 40.45 g/l, 7.9, 30.29 oC and 86.28 h for substrate concentration, pH, temperature and HRT respectively and hydrogen yield of 57.73 ml H2/g TVS. The quadratic polynomial models from the mixture and box-behnken design had a coefficient of determination (R2) of 0.94 and 0.79 respectively, suggesting that the models were adequate to navigate the optimization space. The feasibility of a large-scale biohydrogen fermentation process was studied using the optimized operational setpoints. A semi-pilot scale biohydrogen fermentation process was carried out in 10 L bioreactor and the potential of generating bioelectricity from the process effluents was further assessed using a two-chambered Microbial Fuel Cell (MFC) process. The maximum hydrogen fraction of 46.7% and hydrogen yield of 246.93 ml H2/g TVS were obtained from the semi-pilot process. The maximum electrical power and current densities of 0.21 W/m2 and 0.74 A/m2 respectively were recorded at 500 Ω and the chemical oxygen demand (COD) removal efficiency of 50.1% was achieved from the MFC process. This study has highlighted the feasibility of applying agricultural and municipal wastes for large-scale microbial production of hydrogen, with a simultaneous generation of bioelectricity from the process effluents. Furthermore, the potential of generating an economical feasible biohydrogen production process from these waste materials was demonstrated in this work. Keywords: Biohydrogen production, Organic Fraction of Solid Municipal Waste (OFSMW), Modelling and optimization, Fermentation process, Renewable energy, Bioenergy / Thesis (M.Sc.)-University of KwaZulu-Natal, Pietermaritzburg, 2013.

Submitted in fulfillment of the requirements Of the degree of Masters in applied science: Biotechnology, Durban University of Technology, Durban, South Africa, 2016. / Anaerobic dark fermentation is an efficient biological process to produce hydrogen from waste material. In South Africa, this technology has not been explored adequately to extract energy from biological wastes. Within the KwaZulu Natal region of South Africa, the sugar industry is a prominent venture that produces mass quantities of sugarcane bagasse amongst other waste products. This by-product can be an ideal source of substrate for biohydrogen generation. In this study, sugarcane bagasse was used as the main substrate for biohydrogen production by anaerobic fermentation using sewage sludge as the inoculum. Different pre-treatment methods were employed to maximize the release of fermentable sugars from the lignocellulosic biomass. Among the different pre-treatment methods employed, the maximum sugar yield (294.4 mg/g) was achieved with 0.25% H2SO4 for 60 minutes at 121°C. Prior to inoculation, the sewage sludge was also subjected to thermal pre-treatment to eliminate methanogens. Thermal pre-treatment of inoculum sludge for 30 min was effective in eliminating methanogens. Fluorescence in situ hybridization was used to positively identify the hydrogen producing bacteria present before and after treatment. The pre-treated substrate and inoculum was integrated into a dark fermentation process to further optimize the effect of pH, substrate to biomass, iron and magnetite nanoparticles on hydrogen production. The maximum hydrogen production (1.2 mol/mol glucose) was achieved at a pH range of 5-6, a substrate to biomass ratio of 3.5, and iron and magnetite nanoparticle concentration of 200 mg/L. Microbial analysis using quantitative polymerase chain reaction has confirmed the dominance of Clostridium spp. in the reactor. The highest hydrogenase gene activity (number of copies of hydrogenase gene expression/ng DNA) was recorded in the reactor supplemented with magnetite nanoparticles with lowest being in the raw sludge. There was a direct positive correlation between the hydrogenase gene copy number and the hydrogen yield obtained at different reactor conditions. Scanning electron microscopy was a useful to visually analyse the interaction of microorganisms with activated sludge. This study highlights the significance of anaerobic microorganisms from waste sludge being able to utilize agricultural waste material to produce biohydrogen which could be further scaled up for continuous hydrogen production. In addition, statistical tools used to predict the possible sugar (Design of experiments) and hydrogen yields (Gompertz model) produced would be helpful in saving time during full-scale operation of biohydrogen producing reactors. / M

A dissertation submitted to the Faculty of Engineering and the Built Environment, University of the Witwatersrand, Johannesburg, in fulfillment of the requirements for the degree of Doctor of Philosophy in Engineering, October 2017 / The dwindling fossil reserves coupled with environmental pollution necessitate the search for clean and sustainable energy resources. Biohydrogen is emerging as a suitable alternative to fossil fuels and has received considerable attention in recent years due to its economic, social, and environmental benefits. However, the industrial application of biohydrogen has been hindered by low yield. Therefore, development of novel techniques to enhance the yield is of immense importance towards large-scale production of biohydrogen. Thus, this research effort explored various options to enhance the yield of biohydrogen during dark fermentation process. Some options explored included (i) the utilization of feedstocks from the agricultural, industrial and municipal sectors, (ii) parametric optimization of biohydrogen production, (iii) investigation of biohydrogen production using metal ions and nitrogen gas sparging, and (iv) assessing the feasibility of biohydrogen scale-up study to pave the way for pilot-scale development. Solid biowaste feedstocks consisting of apple, bread, brewery residue, cabbage, corn-cob, mango, mealie-pap, pear, potato, and sugarcane were investigated for dark fermentative biohydrogen production using anaerobic mixed sludge. The experimental results showed that substrates which are rich in carbohydrates are suitable for dark fermentative biohydrogen-producing bacteria. Consequently, a maximum biohydrogen fraction of 43.98, 40.32 and 38.12% with a corresponding cumulative biohydrogen yield of 278.36, 238.32 and 215.69 mL H2/g total volatile solids (TVS) was obtained using potato, cabbage, and brewery wastes, respectively. Based on these results, potato waste was chosen as a suitable substrate for subsequent biohydrogen production studies. Parametric optimization was carried out on biohydrogen production via dark fermentation using potato waste as the substrate. Effects of operating variables such as pH, temperature, fermentation time, and substrate concentration were investigated via response surface methodology (RSM) approach using a two-level-four factor (24) central composite design (CCD). The obtained predictive model (statistical model) was used to explain the main and interaction effects of the considered variables on biohydrogen production. In addition, the model was employed in the optimization of the operating conditions. Consequently, a secondorder polynomial regression with a coefficient of determination (R2) of 0.99 was obtained and used in the explanation and optimization of operating variables. The optimum operating conditions for biohydrogen production were 39.56 g/L, 5.56, 37.87 oC and 82.58 h for potato waste concentration, pH, temperature and fermentation time, respectively, with a corresponding biohydrogen yield of 68.54 mL H2/g TVS. These results were then validated experimentally and a high biohydrogen yield of 79.43 mL H2/g TVS indicating a 15.9% increase was obtained. Furthermore, the optimized fermentation conditions were applied in the scale-up study of biohydrogen production that employed anaerobic mixed bacteria (sludge) which was immobilized in calcium alginate beads. A biohydrogen fraction of 56.38% with a concomitant yield of 298.11 mL H2/g TVS was achieved from the scale-up study. The research also investigated the influence of metal ions (Fe2+, Ca2+, Mg2+ and Ni2+) on biohydrogen production from suspended and immobilized cells of anaerobic mixed sludge using the established optimal operating conditions. A maximum biohydrogen fraction of 45.21% and a corresponding yield of 292.8 mL H2/g TVS was achieved in fermentation using Fe2+ (1000 mg/L) and immobilized cells. The yield was 1.3 times higher than that of suspended cultures. The effect of nitrogen gas sparging on biohydrogen conversion efficiency (via suspended and immobilized cells) was studied as well. Cell immobilization and nitrogen gas sparging were effective for biohydrogen production enhancement. A maximum biohydrogen fraction of 56.98% corresponding to a biohydrogen yield of 294.83 mL H2/g TVS was obtained in a batch process using nitrogen gas sparging with immobilized cultures. The yield was 1.8 and 2.5 times higher than that of nitrogen gas sparged and non-sparged suspended cell system, respectively. Understanding the functional role of microorganisms that actively participate in dark fermentation process could provide in-depth information for the metabolic enhancement of biohydrogen-producing pathways. Therefore, the microbial composition in the fermentation medium of the optimal substrate (potato waste) was examined using PCR-based 16S rRNA approach. Microbial inventory analysis confirmed the presence of Clostridium species which are the dominant biohydrogen-producing bacteria. The results obtained from this research demonstrated the potential of producing biohydrogen using South African solid biowaste effluents. These feedstocks are advantageous in biohydrogen production because they are highly accessible, rich in nutritional content, and cause huge environmental concerns. Furthermore, optimization techniques using these feedstocks will play a pivotal role towards large-scale production of biohydrogen by increasing throughput and reducing the substrate costs which accounts for approximately 60% of the overall costs. The findings from this research also provide a solid basis for further scale-up and techno-economic studies. Such studies are necessary to evaluate the competitiveness of this technology with the traditional processes of hydrogen production. In summary, the findings from this research effort have been communicated to researchers in the area of biohydrogen process development in the form of peer-reviewed international scientific publications and conference proceedings, and could provide a platform for developing an economic biohydrogen scaled-up process. / CK2018