Comparative analysis of biohydrogen producing bacterial consortia in three thermophilic anaerobic fluidised bed bioreactors

Sebola, Keneilwe Mmule
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Global warming has stimulated research into alternative energy carries and fuels. Hydrogen is one of these alternative fuels that are recognized as a promising future energy source. Historically, it is produced by water electrolysis and the gasification of coal. Hydrogen is a natural though transient by-product of several microbial driven biochemical reactions, including anaerobic digestion and fermentation. Microorganisms degrade complex molecules to produce butyrate and alcohols with CO2 and hydrogen as the only by- products. Hydrogen produced by microorganisms is known as biohydrogen. This study aimed to identify biohydrogen- producing bacteria in three Anaerobic Fluidised Bed Bioreactors (AFBRs), which are capable of producing hydrogen under anaerobic and thermophilic conditions, by using PCR-DGGE analysis of the 16 S rDNA genes. Sewage sludge from anaerobic digester and cow- dung were used as inoculum to isolate potential H2- producing organisms. The operational conditions were manipulated to removing mesophilic bacteria and non- spore forming bacteria by gradually increasing the temperature to thermophilic conditions and the pH maintained at acid conditions to allow acidotolerant bacteria to survive. The bioreactors were operated for a period of forty days for each research cycle. Bioreactor One was operated at 55°C for eight days and then the temperature kept at a constant 65°C for the remainder of the research cycle. Bioreactor Two was operated for 8 days at each temperature ranging from 45°C - 65°C, with increments of 5 °C. Bioreactor Three was operated at each temperature for ten days ranging from 55- 70°C, also with increments of 5°C. Samples for microbial community identification were taken at 55°C and 65°C. The bacterial morphologies and structural properties were evaluated by examining the hydrogen- producing granules, isolated at 65°C, using scanning electron microscopy. Species of the families Bacillus, Enterobacteria, Actinomyces, Clostridium and Veillonella were identified. Clostridium thermopalmarium, Bacillus coagulans and Bacillus thermoamylovorans were the culturable species at the desired operational temperature of 65°C. C. thermopalmarium and B. coagulans are major H2 producers with theoretical values of 4mol H2/ g glucose and 3mol H2/ mol sugar, respectively. Sewage sludge is the best source of biohydrogen producing bacteria in comparison to cow dung as an inoculum in AFBRs. Bioreactor 1 with a constant operational temperature of 65°C and at a HRT of 6.5 allowed for the desired Clostridium sp. to be the predominant H2 producer.
A research report submitted to the Faculty of Health Sciences, University of the Witwatersrand, in partial fulfilment of the requirements for the degree of Master of Science in Medicine (Pharmaceutical Affairs). Johannesburg 2012