Toxicity and Inhibition 

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Ryan Hennessy
Non Viable Red Fluorescent

Toxicity and Inhibition are two important but distinctly different concepts in biological wastewater treatment processes. While both are viewed as negative, it is important to recognize the differences in these terms. The following will start with an explanation on toxicity and go on to explain the areas of overlap as well as the distinct differences in these terms. 


Dr. Michael Richard defined toxicity as events in which the oxygen uptake rate (OUR) of the bacteria when fed increasing amounts of a substance, reduced OUR (oxygen uptake rate) values below the “unfed”/ baseline value. For example, if a stable RAS (return activated sludge) sample were collected and DO (dissolved oxygen) depletion over time were measured, this may have an OUR value of around 10 mg/L/hr. When the RAS sample is fed substrate, increases in oxygen uptake rate should increase in correspondence with the amount of added substrate (food). As an example: adding 2 mLs of substrate may result in an increase of OUR from 10 to 15 mg/L/hr., 5 mLs addition may increase the OUR to 25 mg/L/hr., and 10 mLs addition may increase the OUR value to 40 mg/L/hr. representing a “normal/healthy” curve.  In an event in which toxicity occurs, at a certain value of added substrate, the OUR value decreases below the baseline value (in this case 10 mg/L/hr. for an unfed RAS sample). 

In practice, with respirometry testing it is common to see normal correlations of OUR with various amounts of fed substrate until a toxicity “threshold” is reached. There is commonly an initial increase in OUR values due to stress, followed by an often-sharp decline in the OUR value once the concentration of the substance added reaches a toxicity threshold value and bacteria begin to die (dead bacteria don’t use oxygen).

Toxicity may be measured in a wholistic approach (i.e., OUR testing) or may also be more specific to microbes that are impacted by an event (more challenging to test for). It is important to recognize that biological wastewater treatment processes encompass a highly competitive environment for growth, and it is sometimes possible that if a microbe has the capabilities of “filling the void” and competing in an environment that may be toxic to the former microbe, the corresponding impacts on biological treatment performance may be variable. 

Toxic Substances 

Toxicity occurs when the crippling of the given microbe’s enzyme functionality results in death. The most significant factor of any substance is its concentration and impact on the toxicity threshold of the microbial community in the treatment system. Substances recognized as “toxic” tend to have lower concentrations in which they begin to create toxic environments. Most substances are toxic at certain concentrations, however “safer” substances generally are not found in concentrations in which they may have negative impact. Substances most often correlated with toxicity include heavy metals (i.e., chromium, mercury, lead, arsenic), various organic compounds (i.e., solvents, phenols, herbicides, hydrocarbons), petroleum products, pharmaceuticals, chlorinated compounds, and other oxidizers/cleaning agents such as quaternary ammonium compounds and peracetic acid. 

Inhibition versus Toxicity 

Inhibition overlaps with toxicity as it often begins to occur prior to a toxic threshold concentration. Inhibition disrupts the enzyme activity of microbes and delays the rate of treatment. In respirometry testing a curve showing inhibition would show that as a certain amount of substrate is added, the OUR value begins to decrease rather than increase. When the decrease of the OUR value exceeds the initial “unfed/baseline” value of the RAS sample, inhibition technically switches to toxicity when following Dr. Richard’s protocols. Inhibition may have varying degrees of impact on total treatment depending upon how much time is available for the microbes to recover after inhibition occurs.

Most microbes in wastewater treatment processes are responsible for various functions which rely on the ability of their enzymes to work optimally. When inhibition is present, the microbe doesn’t actually die, rather it decreases or in some instances completely stops its functionality until its enzymes are able to perform again. 

Operational Control Strategies

Under most circumstances decreasing the WAS (waste activated sludge) rate allows for a greater “buffer” for stressful events so that even if a certain percentage of bacteria are compromised, enough bacteria may still be functional for maintaining treatment performance. Many plants have success running the MLSS (mixed liquor suspended solids) concentration and the SRT (sludge retention time) values on the higher ends to allow this buffering function. The degree of flexibility for higher SRT and MLSS values generally depends on the hydraulic and solids loading capabilities of the final clarifier (s) as well as the settling characteristics of the mixed liquor (i.e., SVI- sludge volume index). 

Because concentration of a toxin is the most significant parameter, if a known toxic steam is segregated, there are times in which it may bled in slowly at a rate that is safe and under the toxic threshold. Dr. Richard recommended respirometry testing for these decisions, and not to accept any waste that exceeded a 10% reduction in OUR values at any applied rate. In some instances, separation as well as neutralizing the toxin may be achievable (case specific). 

I recommend a microscopic evaluation immediately after the time of an event in question to gauge the overall health of the biomass. Generally, when accessing the overall health and conditions of the bacteria, input from the consultant helps provide direction, while the experience of the operator and their familiarity with the plant is extremely valuable to determine at which degree to implement operational changes. When a notable toxic event has occurred, usually a form of reduced wasting (but still some degree of wasting to rid the system of dead bacteria) yields success provided that the toxic substance doesn’t continue to enter the treatment plant. Dilution (i.e., higher RAS rates or higher flows of known healthy substrate) is also often beneficial. 

Quaternary Ammonium Compounds and “Legacy” Toxicity and Inhibition

While under most circumstances increasing the MLSS and SRT values tends to be successful, a notable exception is when quaternary ammonium compounds(QAC) are present. Quaternary ammonium compounds may have acute as well as chronic (long term) impacts on treatment performance. Chronic inhibition and toxicity often occur when the concentration of QAC compounds bound within the biosolids exceeds 10 mg/. Because of this “legacy” toxicity impact, there are times in which aggressive wasting corresponding with re-seeding of a healthy biomass are warranted so the quats are ultimately removed from the system. Note that aggressive wasting must usually be done in conjunction with re-seeding of adequate bacteria to achieve the growth of desirable bacteria, especially in industrial wastewater treatment processes. 

 As a rule, it is recommended that any treatment plant that receives quaternary ammonium compounds have these compounds neutralized prior to entering biological wastewater treatment. My direct expertise is in microscopy and process control troubleshooting; however I may be contacted directly as I often refer clients to competent professionals that specialize in these areas related to quaternary ammonium compounds.   


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