Understanding how temperature and substrate influence wastewater nitrification


By Nick Szoke

Good science dictates that the protection of aquatic life needs to consider the most sensitive life stages of all species at risk. As such, depending on the aquatic life diversity and species present in the receiving water body, monthly or seasonal maximum ammonia loading and/or concentration limits are commonly set by the regulatory authority.

Low limits are typically set for spring conditions, the most sensitive time of the year, to protect fish during spawning and recently hatched fish. This also tends to correspond to the spring freshet that results in cold extraneous surface water entering a sewer system, depressing the influent water temperature and diluting the constituent concentrations. Cold dilute influent creates a challenge for operators trying to maintain the level of nitrification required to meet low NH3 effluent limits during this period.

Wastewater nitrification is a two-step process that requires an aerobic environment and sufficient alkalinity to support the growth of chemoautotrophic nitrifier populations. It is important to note that total ammonia nitrogen (TAN) is the sum of un-ionized ammonia (NH3) plus ammonium (NH4+). Total nitrite nitrogen (TNN) is the sum of nitrite (NO2ˉ) and nitrous acid (HNO2ˉ). This clarification is very important since the equilibriums between NH3 ←→ NH4+ and NO2ˉ ←→ HNO2ˉ are a function of temperature and pH.

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Un-ionized ammonia (NH3), often referred to as free ammonia (FA), is a form toxic to aquatic life if present in large enough concentrations. Moreover, HNO2, often referred to as free nitrous acid (FNA), is highly toxic to living organisms in very small quantities. NH4+ is the fraction of TAN used by ammonia-oxidizing bacteria (AOB) to convert it to nitrite (NO2ˉ).

The process of nitritation (NH4+ → NO2ˉ) consumes alkalinity, which can drive down pH and increase the concentration HNO2ˉ in the process to a point that can inhibit the nitrification process. This is a self-limiting process, if alkalinity is in short supply. In some cases, it is possible to nitrify in low pH environments (above 6.5), if sufficient alkalinity exists, because the environment acts as a natural selector and changes the bacterial community structure. Essentially, this is survival of the fittest.

A rapid drop in pH will result in a very harsh condition, since there is inadequate time for the community to adapt and acclimate to the new environment. For normal conditions, it is important to keep pH above 7.0 and in the optimal range of 7.5 to 8.5 pH units. Nitrite-oxidizing bacteria (NOB) convert NO2ˉto nitrate (NO3ˉ) but consume alkalinity to a much smaller degree.

There are many different biological nutrient removal (BNR) processes that include nitrification (Johannesburg, Bardenpho, Step-feed, Westside, Modified Ludzack-Ettinger, University of Cape Town, etc.) with or without denitrification and phosphorus (chemical or biological) removal. The BNR treatment process normally includes a secondary clarifier, which is a critical unit operation in an activated sludge (AS) process. The AS BNR process will be configured differently (i.e., aerobic, anoxic, and anaerobic zones) depending on the treatment process employed to achieve the desired effluent quality.

Denitrification requires an anoxic zone (oxygen only in the form of NOx), while biological phosphorus removal requires an anaerobic zone (devoid of all forms of oxygen) and a source of volatile fatty acids (VFA). Chemical phosphorus removal via a metal salt can be done by adding the chemical prior to the primary or secondary clarifiers, but can reduce the pH of the wastewater stream.

Biological nitrification is performed by two chemoautotrophic nitrifying microorganisms in sequence, AOB then NOB. The anabolic and catabolic processes of AOBs and NOBs are influenced by a host of factors: Growth rate (µmax) and solids retention time (SRT); decay rate (b); half saturation (Ks – also referred to as affinity); substrate concentrations (NH3, NO2, HNO2, O2, alkalinity, P, etc.); temperature (T) and Arrhenius (Ѳ); pH; alkalinity in the form of CO2; oxygen concentration; C/N/P ratio; microbial community composition; light; inhibitory or toxic compounds; hydraulic retention time (HRT) and process configuration.

Temperature influence on the nitrification processes is an important factor in the design and operation of a BNR treatment process. In an AS process, the influence of temperature on process kinetics can be expressed by the following equation:

Rate adjustment coefficient, kT = k20 ∙ Ѳ(T-20)


  • k20 is the reaction rate at a reference temperature of 20°C
  • Ѳ is the temperature Arrhenius activity coefficient (unitless)
  • T is temperature (°C)

The maximum specific growth rate of nitrifiers (µmax) and maximum decay rate (bmax) combine together to establish the net growth rate of nitrifiers (µnet) at a reference temperature of 20°C.

  • µnet = (µmax,20 ∙ Ѳµ(T-20)) – (bmax,20 ∙ Ѳb(T-20))

Since nitrification is a two-step process, the kinetic rates influenced must be adjusted independently to properly simulate the biochemical conversion of ammonium to nitrite, and from nitrite to nitrate. The difference in growth rates and environmental conditions can result in nitrite accumulation in the system and prevent full nitrification from occurring. This condition is referred to as nitrite-lock and requires a closer inspection of the cause (e.g., low pH, low DO, low alkalinity, temperature, startup lag, etc.) to properly address the situation.

  • AOB, µAOB,max,20 = 0.90 (d-1), ѲAOB,µ = 1.072, KAOB,s = 0.70 (mg/L-N), bAOB,max,20 = 0.15 (d-1), ѲAOB,b = 1.029
  • NOB, µNOB,max,20 = 0.70 (d-1), ѲNOB,µ = 1.060, KNOB,s = 0.10 (mg/L-N), bNOB,max,20 = 0.15 (d-1), ѲNOB,b = 1.029

The following provides an example calculation to illustrate the impact of cold influent on the nitrification process. A summer influent can have a temperature of 20°C, while influent temperatures can be depressed to 8°C during the spring freshet if too much extraneous flow enters the sewer system.

  • µnet, AOB, 20°C = (0.9 ∙ 1.072(20-20)) – (0.15 ∙ 1.029(20-20)) = 0.75 d-1
  • µnet, AOB, 8°C = (0.9 ∙ 1.072(8-20)) – (0.15 ∙ 1.029(8-20)) = 0.28 d-1

The above calculation highlights that it takes 2.6 times longer to grow AOBs in a colder environment.

The half saturation constant (Ks) used in switching functions is based on an empirical Monod type relationship to adjust the growth response based on substrate (S) concentration, as given below.

  • µactual = µnet [S/(S+Ks)]

Assuming no TAN limitation, nitrification requires an aerobic environment (i.e., DO ≥ 2 mg/L) with sufficient alkalinity to sustain the biochemical process. To account for more than one limiting substrate (e.g., oxygen, alkalinity, nutrients), switch functions for each substrate [S/(Ks + S)] are multiplied together to account for the synergistic effect on growth rate. Specifically, as the substrate concentration decreases, the growth rate decreases, and becomes very pronounced below the Ks value.

Dilution of influent due to extraneous inflow into the sewer system can negatively impact the nitrifier biomass that can be grown in the bioreactors due to the diminished growth rate. A higher concentration at the start of the bioreactor, the faster the kinetics. Starting at a lower initial concentration near the Ks value will significantly hinder the growth rate.

The aerobic solids retention time (SRTa) required to grow nitrifiers is the inverse of the actual growth rate, SRTa = 1/µactual. Factoring in the reduction in nitrifier growth due to colder influent and the dilute substrate concentration from extraneous inflows, a longer SRT is required to achieve the desired degree of nitrification to meet effluent ammonia requirements. Simply put, a longer SRT will result in more solids inventory and higher mixed liquor suspended solids (MLSS) in the bioreactors.

Too high a MLSS in the bioreactors can exceed the safe solids loading rate (SLR) of the final clarifiers and cause non-compliance with final effluent requirements. There is also the risk of possible failure of the nitrification process due to an abrupt change like washout (i.e., SRTa below 1/µactual) and the inability to achieve the final effluent ammonia concentrations due to the loss of nitrifier mass.

Maintaining a larger inventory of AS in the final clarifier to support nitrification can, in certain cases, cause bulking or rising sludge to occur. This is due to the fact that the sludge age or mean cell residence time (MCRT), which is different than the SRTa, can change in the community structure (i.e., fair excessive filamentous growth) and/or cause the final clarifier to behave as a bioreactor, resulting in denitrification and the formation of nitrogen gas bubbles. As such, bulking or rising sludge can cause a deterioration in final effluent quality. Incomplete denitrification in mainstream process during stressed conditions resulting from cold dilute flows can also lead to rising sludge in the final clarifiers.

Stantec has designed and upgraded BNR wastewater treatment plants across North America to meet the challenging conditions associated with cold weather nitrification. As flows and loads increase to these northern wastewater treatment plants, the demands placed on the nitrification process will increase. The design and operation of the nitrification process in a BNR system requires careful attention to several parameters and environmental conditions. Simply increasing the SRTa to compensate for cold dilute inflows can lead to other undesirable conditions if the system was not sized to handle the seasonal episodes of cold dilute flows.

A high sludge volume index (SVI) is an indicator that the capacity of the treatment process is showing signs of stress, and a good indicator for a call to action. A review of the treatment process capacity will help to identify modifications needed to successfully achieve ammonia effluent compliance requirements during this challenging period.

Nick Szoke is with Stantec. This article appears in ES&E Magazine’s April 2019 issue.


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