**4.1 Methanogenic anaerobic digestion**

The anaerobic digestion process is a biological process carried out by three different groups of microorganisms (hydrolytics, acetogenics and methanogenics) (Gujer & Zehnder, 1983)) which transform organic matter to obtain 90% biogas [a mixture CH4/CO2 (≈65%-35%)] and only 10% excess sludge. Biogas has a high calorific value (5000-6000 kcal m-3) and can be used for electricity or heat.

The main advantages of the anaerobic process compared with other types of treatment are (Van Lier, 2007):


Methanogenic anaerobic digestion of organic material has been performed for about a century. Therefore, the food web of anaerobic digestion is reasonably well understood (Figure 2).

Anaerobic digestion of biodegradable wastes involves a large spectrum of bacteria of which three main groups can be distinguished. The first group comprises fermenting bacteria which perform **hydrolysis and acidogenesis** (e.g. *Clostridium butyricum, Propionibacterium*). This involves the action of exo-enzymes to hydrolyze matter such as proteins, fats and carbohydrates into smaller units which can then enter the cells to undergo an oxidationreduction process resulting in the formation of volatile fatty acids (VFA) and some carbon dioxide and hydrogen. The fermenting bacteria are usually designated as an acidifying or acidogenic population because they produce VFA.

Olive Oil Mill Waste Treatment: Improving the Sustainability

expected from **sulfate reducing bacteria.** 

**4.2 Biological hydrogen production** 

promising (Chang et al., 2002).

remains a great challenge.

1997).

2007)

of the Olive Oil Industry with Anaerobic Digestion Technology 281

methylamines are of minor importance in most anaerobic digestion processes. In addition to these three main groups, hydrogen consuming acetogenic bacteria are always present in small numbers in an anaerobic digester. They produce acetic acid from carbon dioxide and

The synthesis of propionic acid from acetic acid, as well as the production of longer chain VFA, occur to a limited extent in anaerobic digestion. Competition for hydrogen can also be

In conventional applications, the acid- and methane-forming microorganisms are kept together inside the reactor system with a delicate balance between these two groups of organisms, because they differ greatly in terms of physiology, nutritional needs, growth kinetics and their sensitivity to environmental conditions. Problems encountered with stability and control in conventional design applications have led researchers to new solutions such as the physical separation of acid-formers and methane-formers in two separate reactors. Optimum environmental conditions for each group of organism is provided separately to enhance the overall process stability and control (Cha & Noike,

Hydrogen is a clean, recyclable, and efficient energy carrier. The possibility of converting hydrogen into electricity via fuel cells makes the application of hydrogen energy very

Hydrogen production via dark fermentation is a special type of anaerobic digestion consisting of only hydrolysis and acidogenesis. It leads to the production of hydrogen, carbon dioxide and some simple organic compounds [VFA and alcohols]. These readily degradable organic compounds can be used for further methane production. (Bartacek et al.,

Much interest has recently been expressed in the biological production of hydrogen from waste streams by dark fermentation. Biological hydrogen production shares many common features with methanogenic anaerobic digestion, especially the relative ease with which the

From hydrogen-producing mixed cultures, a wide range of species have been isolated, more specically from the genera Clostridium (*Clostridium pasteurianum, Clostridium saccharobutylicum, C. butyricum*), Enterobacter (*E. aerogenes*) and Bacillus under mesophilic conditions; and from the genera Thermoanaerobacterium (*Thermoanaerobacterium thermosaccharolyticum*), and Caldicellulosiruptor (*Caldicellulosiruptor saccharolyticus*, *C. thermocellum, Bacillus thermozeamaize* ) under thermophilic or extremophilic conditions.

However, the low efficiency of the hydrogen production process remains the main limiting factor. Much research will be needed to be carried out to reach hydrogen yields comparable with the theoretical efficiency maximum. Although a relatively high efficiency has been reached using pure substrates, the low hydrogen yield with complex (real) substrates

two gaseous products can be separated from the treated waste.

hydrogen and, therefore, compete for hydrogen with the methanogenic archaea.

Fig. 1. Experimental laboratory scale anaerobic reactors (A: reactor, B: pH-meter, C: gasometer with NaOH for methane measurement, D: water bath for temperature control).

Fig. 2. Diagram of the different steps of anaerobic digestion

**Acetogenic bacteria** (e.g. *Clostridium aceticum, Acetobacterium woodii)* constitute the second group and are responsible for breaking down the products of the acidification step to acetic acid. In addition, hydrogen and carbon dioxide are also produced during acetogenesis.

The third group involves methanogenic Archaea (e.g. *Methanobrevibacter smithii, Methanobacterium thermoautotrophicum, Methanosarcina barkerii, Methanotrix soebugenii)* convert acetic acid or carbon dioxide and hydrogen into methane. Other possible methanogenic substrates such as formic acid, methanol, carbon monoxide, and

**B**

**A**

**D**

Fig. 1. Experimental laboratory scale anaerobic reactors (A: reactor, B: pH-meter, C: gasometer with NaOH for methane measurement, D: water bath for temperature control).

**C**

Fig. 2. Diagram of the different steps of anaerobic digestion

**Acetogenic bacteria** (e.g. *Clostridium aceticum, Acetobacterium woodii)* constitute the second group and are responsible for breaking down the products of the acidification step to acetic acid. In addition, hydrogen and carbon dioxide are also produced during acetogenesis.

The third group involves methanogenic Archaea (e.g. *Methanobrevibacter smithii, Methanobacterium thermoautotrophicum, Methanosarcina barkerii, Methanotrix soebugenii)* convert acetic acid or carbon dioxide and hydrogen into methane. Other possible methanogenic substrates such as formic acid, methanol, carbon monoxide, and methylamines are of minor importance in most anaerobic digestion processes. In addition to these three main groups, hydrogen consuming acetogenic bacteria are always present in small numbers in an anaerobic digester. They produce acetic acid from carbon dioxide and hydrogen and, therefore, compete for hydrogen with the methanogenic archaea.

The synthesis of propionic acid from acetic acid, as well as the production of longer chain VFA, occur to a limited extent in anaerobic digestion. Competition for hydrogen can also be expected from **sulfate reducing bacteria.** 

In conventional applications, the acid- and methane-forming microorganisms are kept together inside the reactor system with a delicate balance between these two groups of organisms, because they differ greatly in terms of physiology, nutritional needs, growth kinetics and their sensitivity to environmental conditions. Problems encountered with stability and control in conventional design applications have led researchers to new solutions such as the physical separation of acid-formers and methane-formers in two separate reactors. Optimum environmental conditions for each group of organism is provided separately to enhance the overall process stability and control (Cha & Noike, 1997).
