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For many years, sewage sludge has been processed for methane production in anaerobic digestion reactors at wastewater treatment plants around the world. Sewage sludge is produced in large quantities and is rich in biodegradable organic materials, from which sugars (e.g., glucose) can be produced, recovered and used as a substrate to support hydrogen production through the Dark Fermentation (DF) process. DF is one of several methods used for bio-hydrogen production, whereby fermentative bacteria are used to hydrolyse organic substrates to produce hydrogen gas. Carbohydrates (sugars) is one of the main fermentable substrates for hydrogen production, and they are considered the most favourable substrate for fermentative bacteria (e.g., Clostridium bacteria). Although sewage sludge is rich in organic materials, still the complexity of its structure and low carbon/nitrogen ratio limits the bio-hydrogen production via DF processes. Therefore, this paper addresses the impact of Enzymatic Hydrolysis (EH) as a pre-treatment of sewage sludge on enhancing the biodegradability and glucose content in sewage sludge. The result shows that using the EH process as pre-treatment for sewage sludge, enhanced the glucose content in sewage sludge and converted some of the macro sewage flocs to easy digestible micro flocs (glucose). Therefore, the substrate being more favourable and easier to digest by bacteria in the DF reactor, enhanced the production of hydrogen and VFAs. More research needs to be done to find the optimum enzyme dosage, initial substrate concentration and operation temperature (especially when the enzyme is used inside the DF reactor).

Hydrogen gas (H2) is an energy carrier that does not generate carbon dioxide emissions during combustion, but several processes in use for its production demand high energy inputs associated with fossil fuels and greenhouse emissions. Biological processes, such as dark fermentation (DF), have the potential to remove the dependency on fossil fuels in H2 production. DF is a process that encourages fermentative bacteria to ferment organic substrates to produce H2 as a truly clean energy carrier, but its success depends on removing the presence of competing H2−consuming microorganisms in the inoculum consortia. This paper addresses a strategy to enhance H2 production from different types of substrates by testing inoculum pre-treatment processes to inactivate H2−consuming bacteria, including acid-shock (pH 3), basic-shock (pH 10) and heat-shock (115 °C) methods. Digestate from anaerobic digesters processing sewage sludge was used to produce pre-treated inocula, which were subsequently tested in a batch bio-H2 potential (BHP) test using glucose as a substrate. The results show that heat-shock pre-treatment was the best method, reporting a H2 yield of 191.8 mL-H2/gVS added (the untreated inoculum reported 170.91 mL-H2/gVS added). Glucose conversion data show a high concentration of butyric acid in both treated and untreated inocula during BHP tests, which indicate that the butyrate pathway for H2 production was dominant; shifting this to the formate route could further enhance net H2 production. A standardised inoculum-conditioning method can help to consistently assess the biohydrogen potential of suitable feedstock for DF and maximise H2 yields.