Anaerobic treatment processes require the presence of a diverse closely dependent group of bacteria to bring about the complete conversion of complex mixtures of substrates to methane gas. It is puzzling that single species of bacteria have not evolved to convert at least simple substrates such as carbohydrates, amino acids, or fatty acids all the way to methane [26].

Conventional phase and high-rate two-phase anaerobic digestion processes have frequently been employed in order to treat both soluble and solid types of domestic and industrial wastes. The most significant outcome of anaerobic digestion processes is that they generate energy in the form of biogas namely, methane and hydrogen. Therefore, due to current imperative environmental issues such as global warming, ozone depletion, and formation of acid rain, substitution of renewable energy sources produced from biomass, such as methane and hydrogen, produced through anaerobic digestion processes will definitely affect the demand and consumption of fossil-fuel derived energy [27].

The anaerobic treatment is a biological process widely used in treating wastewater. When these have a high organic load, presents itself as the only alternative would be an expensive aerobic treatment, due to the oxygen supply. The anaerobic treatment is characterized by the production of "biogas" consisting mainly of methane (60-80%) and carbon dioxide (40-20%) and capable of being used as fuel for generating thermal energy and / or electric. Furthermore, only a small fraction of COD treated (5-10%) is used to form new bacteria, compared to 50-70% of an aerobic process. However, the slow anaerobic process requires working with high residence times, so it is necessary to design reactors or digesters with a high concentration of microorganisms [28, 29]. Actually is a complex process involving several groups of bacteria, both strictly anaerobic and facultative, which, through a series of stages and in the absence of oxygen, flows mainly in the formation of methane and carbon dioxide. Each stage of the process, described below, is carried out by different groups of bacteria, which must be in perfect balanced. Figure 1 shows a schematic representation of the main conversion processes in anaerobic digestion, suggested by Gujer [30].

(1) Hydrolysis. In this process complex particulate matter is converted into dissolved compounds with a lower molecular weight. The process requires the mediation of exo­enzymes that are excreted by fermentative bacteria. Proteins are degraded via (poly) peptides to amino acids, carbohydrates are transformed into soluble sugars (mono and disaccharides) and lipids are converted to long chain fatty acids and glycerine. In practice, the hydrolysis rate can be limiting for the overall rate of anaerobic digestion. In particular the conversion rate of lipids becomes very low below 18°C.

(2) Acidogenesis. Dissolved compounds, generated in the hydrolysing step, are taken up in the cells of fermentative bacteria and after acidogenesis excreted as simple organic compounds like volatile fatty acids (VFA), alcohols and mineral compounds like CO2, H2, NH3, H2S, etc. Acidogenic fermentation is carried out by a diverse group of bacteria, most of which are obligate anaerobe. However, some are facultative and can also metabolize organic matter via the oxidative pathway. This is important in anaerobic wastewater treatment, since dissolved oxygen (DO) otherwise might become toxic for obligate anaerobic organisms, such as methanogens [31].

(3) Acetogenesis. The products of acidogenesis are converted into the final precursors for methane generation: acetate, hydrogen and carbon dioxide. As indicated in Figure 1, a fraction of approximately 70% of the COD originally present in the influent is converted into acetic acid and the remainder of the electron donor capacity is concentrated in the formed hydrogen. Naturally the generation of highly reduced material like hydrogen must be accompanied by production of oxidized material like CO2.

(4) Methanogenesis. Methanogenesis may be the rate liminting step in the overall digestion process, especially at high temperatures (> 18°C) and when the organic material in the influent is mainly soluble and little hydrolysis is required. Methane is produced from acetate or from the reduction of carbon dioxide by hydrogen using acetotrophic and hydrogenotrophic bacteria, respectively:

Acetotrophic methanogenesis: CH3COOH ^ CH4 + CO2 (3)

Hydrogenotrophic methanogenesis: 4H2 + CO2 ^ CH4 + 2H2O (4)

Different from aerobic treatment where the bacterial mass was modeled as a single bacterial suspension, anaerobic treatment of complex wastewaters, with particulate matter in the influent, is only feasible by the action of a consortium of the four mentioned groups of bacteria that each have their own kinetics and yield coefficients. The bacteria that produce methane from hydrogen and carbon dioxide grow faster than those utilizing acetate that the acetotrophic methanogens usually are rate limiting for the transformation of acidified wastewaters to biogas [32].

The different groups of bacteria involved in the conversion of influent organic matter all exert anabolic and catabolic activity. Hence, parallel to the release of the different fermentation products, new biomass is formed associated with the four conversion processes described above. For convenience, the first three processes often are lumped together and denominated acid fermentation, while the fourth step is referred to as methanogenic fermentation.

The removal of organic matter-COD during the acid fermentation is limited to the release of hydrogen only 30% of the organic matter is converted into methane via the hydrogenotrophic pathway. Hence, a necessary condition for efficient organic matter removal in an anaerobic treatment system is that a sufficient mass of acetotrophic methanogens develops.

Acid fermentation tends to cause a decrease in the pH because of the production of VFA and other intermediates that dissociate and produce protons. As methanogenesis will only develop well at a neutral pH values, instability may arise, if for some reason the rate of acid removal by methane production falls behind the acid production rate: the net production of acid will tend to cause a decrease in pH, and thus may reduce the methanogenic activity further. In practice, this so called "souring" of the anaerobic reactor contents is the most common cause for operational failure of anaerobic treatment systems. The danger of souring can be avoided, by maintaining the proper balance between acid and methanogenic fermentation which in fact means that both the methanogenic digestion capacity and buffer capacity of the system should be sufficiently high [29, 33].