Bioconversion of Waste Biomass to Biohydrogen


The energy carriers and chemicals are exclusively dependent on fossil fuel sources in the present world scenario. The use of fossil fuels not only increases the concentration of greenhouse gases in the atmosphere but also leads to environmental degradation, pollution, global warming, climate change, acid rain, extreme weather patterns, volatility in the crude oil prices and occasional geopolitical tensions between fuel importing and exporting nations (Nanda et al. 2016b; Rana et al. 2018; Rana et al. 2019; Rana et al. 2020; Vakulchuk et al. 2020). With many dependencies on fossil fuel energy sources such as crude oil, gasoline, diesel, coal and natural gas, the major global concern is to seek an alternative energy carrier and vector that can make a paradigm shift in the energy usage from fossil fuels to biofuels for long-term scenarios (Nanda et al. 2015b). Among all biofuel sources, hydrogen has great potentials for use as an outstanding energy carrier and vector. Being a next-generation biofuel, hydrogen can be utilized as a direct fuel, in fuel cells or as a precursor to producing advanced hydrocarbon fuels and chemicals (Nanda et al. 2017b; Okolie et al. 2019; Bhatia et al. 2020; Sarangi and Nanda 2020; Okolie et al. 2020a).

There is an increasing interest in hydrogen generation and utilization as a transportation fuel, as a mitigation strategy to the harmful impact of fossil fuels on the environment. Hydrogen (H2) is often referred to as the “fuel of the future” (Reddy et al. 2020). Having a thermal efficiency of about 120 MJ/kg and flame temperature of 2027°C, hydrogen shows an energy content of several magnitudes higher than that of other conventional hydrocarbon fuels (Nanda et al. 2017c). Hydrogen is an environmental-friendly fuel having versatile applications in energy, fuel, combined heat and power, chemical, metallurgy, fertilizers and other commercial and industrial sectors. Hydrogen is considered as a promising renewable energy source for the sustainable future because its combustion generates massive amounts of heat energy and water.

Hydrogen can be produced using a variety of pathways involving thermochemical technologies (e.g. gasification, pyrolysis and reforming), electrolysis, electrochemical, photochemical, photo-catalytic, photo-electrochemical and microbial (photolysis, photo-fermentation, dark fermentation and microbial electrolysis cells) (Nanda et al. 2017b; Huang et al. 2020; Siang et al. 2020). This chapter describes the microbial technologies for biohydrogen production.

Biological Methods for Hydrogen Production

The production of biohydrogen is considered beneficial as far as ecological and energy requirements are concerned. The biological method for biohydrogen production demonstrates less negative impacts on the environment and low requirement of energy as compared to other sources (Nanda et al. 2017b; Sarangi and Nanda 2020). Lignocellulosic biomass contains major fractions of celluloses and hemicelluloses, which could act as the sources of pentose and hexose sugars for microbial conversion to fuels and chemicals (Nanda et al. 2013; Nanda et al. 2014a; Nanda et al. 2017a; Sarangi and Nanda 2018; Sarangi and Nanda 2019; Sarangi et al. 2020). Before the fermentation process, biomass is required to be pretreated and delignified, to remove the lignin and release the monomeric sugars for fermentation to fuels and value-added chemicals (Nanda et al. 2014b; Nanda et al. 2015a; Fougere et al. 2016). Moreover, lignocellulosic biomasses are low-cost feedstocks and abundantly available globally with a continual supply (Nanda et al. 2016a; Okolie et al. 2020b). The production of biohydrogen from waste biomass has been reported extensively in the literature (Magnusson et al. 2008; Guo et al. 2010; Chen et al. 2012; Han et al. 2012; Moodley and Капа 2015; Kumar et al. 2017). Microbial communities utilize a wide variety of biomass resources for generating hydrogen. The types of microorganisms and feedstock have great roles in biohydrogen production. The biological processes for biohydrogen production include photolysis, dark fermentation, photo-fermentation and microbial electrolysis cells (Holladay et al. 2009).

Biohydrogen production through photo-fermentation is performed by utilizing photosynthetic bacteria through the enzyme nitrogenase system with the help of light energy as well as waste biomass. Purple non-sulfur bacteria help in the photo-fermentation process to utilize the reduced organic acids as a carbon source in the presence of solar light, thereby releasing molecular hydrogen with the help of a nitrogenase enzyme system (Basak and Das 2007). Some light-harvesting pigments like chlorophylls, phycobilins and carotenoids support electrons, protons and oxygen by utilizing sunlight via photo-fermentation (Figure 6.1). The nitrogenase enzyme system helps in the reaction of protons, electrons and nitrogen along with adenosine triphosphate (ATP) to produce hydrogen, ammonia, adenosine diphosphate (ADP) and inorganic phosphates (Pi) (Nanda et al. 2017b). The bacterial photosystem produces two electrons with four ATP molecules by utilizing light energy and biomass to generate hydrogen with the aid of the nitrogenase system.

A simplified illustration of the photo-fermentation process for biohydrogen production

FIGURE 6.1 A simplified illustration of the photo-fermentation process for biohydrogen production

Dark fermentation is considered as a promising method for hydrogen production from waste biomass and specific microorganisms (Kumar et al. 2017; Lukajtis et al. 2018; Sarangi and Nanda 2020). During this method, a mixed gas containing H, and CO, is produced along with other gases like CH4, CO and H,S, which depends on the type of feedstock, microorganisms and process conditions (Datar et al. 2004; Najafpour et al. 2004; Kotsopoulos et al. 2006; Temudo et al. 2007). In dark fermentation, the bacterium converts organic substances like raw biomass, sugars and wastewater to hydrogen. Due to the complete absence of light, this process is regarded as dark fermentation. Moreover, dark fermentation is advantageous over photo-fermentation in requiring smaller bioreactors, less energy and low cost because of the absence of light energy to facilitate microbial growth. Some notable microorganisms like anaerobic bacteria such as Bacillus spp., Enterobacter spp. and Clostridium spp. are utilized for the conversion of cellulosic substrates to biohydrogen (Levin et al. 2004). During the dark fermentation. the bacterium converts glucose to pyruvic acid, thereby producing ATP through the glycolytic pathways. Furthermore, with the utilization of pyruvate ferredoxin oxi- doreductase and hydrogenase, CO, and H, are produced from pyruvic acid (Figure 6.2). During dark fermentation, biohydrogen production generally depends on the degradation of pyruvate to acetyl-CoA and further to acetate, butyrate and ethanol.

Depending on the microorganisms employed and process conditions, different end products and by-products are generated from microbial biohydrogen production. The optimization of fermentation process parameters like sugar content, nutrients (including energy source and carbon source), hydrogen partial pressure, temperature, hydraulic retention time, pH. type of microorganisms used, inoculum pretreatment process, growth medium and cultural conditions can enhance the production of biohydrogen

A simplified illustration of the dark fermentation process for biohydrogen production

FIGURE 6.2 A simplified illustration of the dark fermentation process for biohydrogen production

(Ghimire et al. 2015). Besides, the bioreactor configuration, geometry and mode of operation can also affect biohydrogen production (Show et al. 2011).

Hydrogen production by microbial electrolysis cells is also regarded as a potential method for the utilization of energy as well as protons by microorganisms to convert the organic matter. Not only hydrogen but also various value-added chemicals are generated through this method, thereby establishing it as a promising platform for bioenergy utilization. Different value-added platform chemicals such as methane, formic acid and hydrogen peroxide are also released during this process with the potentials of wastewater treatment (Escapa et al. 2016). The process is similar to microbial fuel cells in having two compartments of anode and cathode separated by a proton exchange membrane (Nanda et al. 2017c; Bhatia et al. 2020). Protons (H+) and electrons (e ) are produced due to the oxidation of organic matter during this process. These are subsequently transferred to the cathode side through the membrane. Hydrogen is produced near the cathode by the reduction of electrons and protons in the presence of a catalyst (Logan et al. 2006).


Biohydrogen generation by microbial biomass is gaining attention considering its environmental friendliness and sustainability. Similarly, biohydrogen from waste biomass and organic residues are regarded as the most promising feedstocks owing to their renewability, abundance, low cost and polysaccharide composition. The exploration of microbial diversities along with the optimization of photo-fermentation and dark fermentation can maximize biohydrogen production. The integration of novel technologies in microbiology, chemical engineering and bioprocess engineering can create a roadmap for efficient and sustainable biohydrogen production leading to energy security, economic sustainability and mitigation of greenhouse gas emissions.


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