Impact of high ammonia concentrations

Ammonia plays a key role in the performance and stability of anaerobic reactors operated on substrate with a high protein content. Ammonia has a positive impact on AD, as bacteria need nitrogen for their cell mass synthesis, they obtain it directly from NH4-N.

In fresh blackwater, most nitrogen is present in the form of urea, a product of protein degradation. Urea (CO(NH2)2) is decomposed by bacteria via the following enzymatic catalyzed reaction (Fidaleo and Laveccio 2003):

1)

CO(NH2)2 + 2 H2O → 2 NH4+ + CO32- (

Ammonia and the buffer capacity

One of the most important parameters affecting reactor stability concerns the buffer capacity of the system. The bicarbonate-ammonia buffer is the primary parameter controlling the pH and process stability. The buffer capacity of a solution is determined from the concentrations of each buffer present, here ammonia and carbonate, their pK values and the pH of the solution. Due to the decomposition of urea, ammonia and carbonate are present in high concentrations in blackwater and therefore the system has a high buffer capacity. Consequently, a dramatic pH-fall below 6 as a critical value hardly occurs (van Velsen and Lettinga 1980). However, the formation of VFA (HAc) decreases the buffer capacity but the formation of NH4+ produced at the same time increases the bicarbonate concentration:

2)

HAc + NH4 HCO3 → NH4 Ac + H2CO3 (

Additional buffer capacity has sulphate contained in the substrate. As sulphate is of importance within AD, its behavior is shortly described here with the following equations:

3)

SO42- + 4 H2 → H2S + 2 H2O + 2 OH- (

4)

SO42- + CH3 COOH → H2S + 2 HCO3- (

The sulphate reduction leads to a COD decrease and an increase of pH and buffer capacity.

Partly the H2S is leaving the system with the biogas flow.

If the buffer capacity is high, the AD process is very stable. If the pH increases due to high ammonia or sulphate concentrations, it subsequently inhibits the methanogenic bacteria and the

declined bacterial activity leads to a VFA accumulation so that a new steady state process will be established. If the pH drop is too drastic, the AD process may irreversibly break down.

Inhibition by ammonia

High ammonia concentrations can cause inhibition of microbiological activity in the AD process.

Ammonia is frequently considered as a cause for digester failure (El Mashad 2003). AD of swine manure as sole substrate was sometimes unsuccessful and the reason was assumed to be the high content of ammonia of more than 4,000 mg/l NH4-N in this waste (van Velsen 1979). An ammonia concentration of 4,000 mg/l NH4-N was shown to be inhibitory during digestion of cattle manure (Angelidaki and Ahring 1994). Ammonia concentrations exceeding 3,000 mg/l NH4-N are toxic regardless of the pH according to van Velsen (1979) and Koster and Lettinga (1984). For unadapted methanogenic cultures, ammonia inhibition has been observed to start at concentration of 1,500 to 2,500 mg/l NH4-N (van Velsen 1979b, Hansen et al. 1998). By adaptation of the biogas process to ammonium, tolerance to more than 4,000 mg/l NH4-N has been demonstrated for swine as well as for cattle manure (Hashimoto 1986, Angelidaki et al.

1993). But still it is not clear, why AD of swine manure with similar NH4-N concentrations is more sensitive to ammonia inhibition than AD of cattle manure (El-Mashad 2003).

Ammonia inhibits predominantly the methanogenesis (Koster and Lettinga 1984, Kroiss 1986, Angelidaki et al. 1993). Acetate utilizing methanogenic bacteria were found to be more sensitive to ammonia than hydrogen consuming ones (Poggi-Varaldo et al. 1997). The active component causing ammonia inhibition is considered to be the free ammonia concentration (Hansen et al.

1998). Two different mechanisms were attributed to ammonia inhibition: Firstly methane synthesizing enzymes are directly inhibited by free ammonia. Secondly the bacteria cell wall is far more permeable to undissociated molecules than to ions. In the cell, free ammonia is rapidly converted to NH4+ changing intracellular pH conditions (Kadam and Boone 1996).

The concentration of the ammonia ion (NH4+) and free ammonia (NH3) are interrelated via the pH and temperature according to the following equation:

5)

NH3 + H2O ↔ NH4+ + OH- (

The higher the pH and the temperature, the higher the free ammonia concentration is. If rising free ammonia concentration starts to inhibit the methanogenic bacteria, VFA concentration increases. This leads to a decrease of the pH and the free ammonia concentration again. In a certain range, the process stabilizes itself (Kroiss 1986).

Acetate utilizing bacteria adapted to ammonia were shown to develop with a free ammonia concentration of up to 700 mg/l NH3-N (Angelidaki et al. 1993), while many lower free ammonia concentrations have been reported for initial inhibition of an unadapted process (Hansen et al 1998). The threshold value of free ammonia is in the range of 80 to 200 mg/l NH3 -N (Table 4).

Table 4: Inhibition thresholds of free ammonia for mesophilic anaerobic digestion

Substrate Free ammonia

concentration pH Reference

Unit mg/l NH₃-N

Potato juice 80 – 150 7.5 Koster and Lettinga 1984 Organic fraction of municipal

solid waste 80 – 100 De Baere et al. 1984 Wastewater from seafood

processing industry 200 Omil et al. 1995 Synthetic wastewater 200 Calli et al. 2005

Several authors found that methane fermentation of high ammonia containing wastewaters is more easily inhibited at thermophilic temperatures than at mesophilic temperatures (Hansen et al.

1998). This coincides with the fact that free ammonia concentration increases with increasing temperature. Furthermore, the biogas process becomes more sensitive towards ammonia when pH increases (Koster 1986) which again increases the concentration of free ammonia.

Hansen et al. (1998) suggested that the interaction between free ammonia, VFA and pH leads to an “inhibited steady state” which is a condition where the process is running stably but with a lower methane rate.

Thus, the ammonia concentration at mesophilic condition has two antagonistic effects, free ammonia causes an inhibition of the process but it also controls the pH and therefore inhibition by accumulation of VFA is avoided and the overall process improved (Vidal et al. 2000). Too high free ammonia concentrations inhibit the process but the extent of the inhibition depends additionally on other factors, such as the adaptation of the microflora and the characteristic of the feedstock.

Prevention of ammonia inhibition

Although the mechanisms of ammonia inhibition are not fully understood, key issues are the free ammonia concentration dependent on pH, and temperature and adaptation conditions. Many researches were dedicated to finding means to improve the performance of AD at high ammonia concentrations, some successful means most relevant for blackwater digestion are the following:

pH control and temperature adjustment: The concentration of free ammonia can be kept lower to prevent ammonia inhibition, if the operation is pH-controlled (Braun et al. 1981). Concerning temperature, mesophilic temperature is apparently more suitable to prevent ammonia inhibition than thermophilic.

Adaptation: Many references mention the importance of bacterial adaptation to ammonia concentrations but it is not clear whether the adaptation is the result of internal changes in the

predominant species of methanogenic bacteria or of a shift in the methanogenic population (El-Mashad 2003). According to Koster (1986) the adaptation of the methanogenic population to high ammonia concentrations is not caused by growing of a new type of bacteria but by slow adaptation of the original population during the period of stagnation of the methane production which lasted in his experiments for around six months.

The adaptation period depends on the applied ammonia concentration and organic loading rate.

The time required for complete acclimation increases with the ammonia concentrations. In un-adapted cultures, free ammonia levels of 150 mg/l NH3-N cause growth inhibition but much higher concentrations can be tolerated by methanogenic bacteria which have undergone a period of gradual adaptation. Adequate adaptation may take two months or even longer (El-Mashad 2003). A linear correlation between the lag phase and the ammonia concentration was found by van Velsen (1981). At ammonia concentrations of 2,000 and 5,000 g/l NH4-N, the lag phase was approximately 20 and 50 days, respectively.

Dilution of the feedstock: By diluting the feedstock, inhibition is prevented by decreasing ammonia concentrations. But this measure increases the required size of the digester and the energy needed to heat the wastewater to operational temperature. Diluting should be applied only in exceptional cases if e.g. the feedstock is too concentrated to be pumped.

Adjusting VS/N or COD/N ratio of the feedstock: More suitable to provide better operational conditions for AD is the addition of substrate that has high organic and low nitrogen content which produces better VS/N ratio for AD. Although the ammonia concentration does not decrease significantly, the process improves in terms of kinetics and becomes more stable.