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Updated: Nov 19, 2020

Adenosine Triphosphate (ATP)—An Introduction

I’ve stated previously (OUR Level 1 Testing page) that whenever I evaluate a biological treatment system there are two sets of tests I always run:

1) Oxygen uptake rate (OUR), and

2) Adenosine triphosphate (ATP) analysis.

When combined, the results of the OUR and ATP analysis provide a very complete picture of conditions in the bioreactor. In this section we'll explore the fundamental use of the ATP test which allows us to make a quick determination of the quantity of living, functioning microorganisms in the bioreactor that actually contribute to the oxidation of organics. Knowing the “true” population of active microorganisms in the bioreactor, we can then accurately calculate the food-to-mass (F:M) ratio. I take the position that most wastewater treatment plants are underestimating the F:M ratio when they use the mixed liquor volatile suspended solids concentration, typically the only parameter they have to estimate their active biomass concentration.

Microbiology image

The mixed liquor suspended solids (MLSS) concentration is a measure of the total solids concentration in the bioreactor. The mixed liquor volatile suspended solids (MLVSS) concentration is a measure of the organic fraction contained in the MLSS. In most biological (activated sludge) treatment systems the MLVSS/MLSS ratio falls within a relatively narrow of 60 to 85 percent as shown in the graph below.

Graphic showing MLVSS/MLSS ratio

The MLVSS value gives a better measure of the portion of the MLSS that is actually contributing to the oxidation of organics. The problem with the MLVSS number is that it includes dead biomass and inert organics as well as living biomass. The very significant value of ATP testing is that it separates the living biomass fraction from the rest of the “organic noise” in the MLVSS sample, as portrayed in the graphic below (modified from the original LuminUltra graphic).

Graphic explaining ATP

Shown in the graphic below are the results of an ATP analysis recently performed at a municipal wastewater plant. On the day of testing the mixed liquor suspended solids (MLSS) concentration was 4,157 mg/L. After combustion, the mixed liquor volatile suspended solids (MLVSS), the assumed concentration of active biomass, was determined to be 3,105 mg/L. The MLVSS/MLSS ratio for these data points is 74.7 percent, a reasonable value. But the far more sophisticated ATP analysis showed the actual concentration of active microorganisms to be just 1,937 mg/L, representing only 46.6% of MLSS in the bioreactor.

AVSS image with example

Overloaded and Unaware

The importance of knowing the true population (concentration) of your biomass cannot be overstated. A common calculation made by wastewater operators is the food-to-mass ratio (F:M ratio). The formula for the F:M ratio is shown below.

"Standard" Food-to-Mass (F:M) Ratio

Food-to-Mass Ratio

The key variable in this formula is the mixed liquor volatile suspended solids (MLVSS) concentration. I am very specifically identifying the MLVSS as being the most significant variable, rather than the BOD (or COD) concentration, because it is the MLVSS, as an indicator of active biomass, that too often (always!) underestimates the fraction of the microorganisms that participate in the oxidation of organics. The result is an underestimation of the F:M ratio and a failure to recognize a biological treatment system that is overloaded, close to being overloaded, or actually overloaded!

The plant referenced in this analysis is a 30 million gallon per day (4,732 m3/hr) conventional, plug flow, activated sludge system. Flow, loading, and aeration volume are shown in the tables below. The recommended range for the F:M ratio in a conventional activated sludge plant is 0.2 to 0.4.

F:M Ratio Comparison

When using the MLVSS number to calculate the F:M ratio the plant appears to be just within the recommended upper limit of 0.4, indicating that the activated sludge system is operating within its design capacity. Believing that the plant is not overloaded makes it that much more difficult to understand why the dissolved oxygen (DO) concentration in the aeration tanks is so low (≤0.5 mg/L), why odors are prevalent (and residents are complaining), and why solids are settling poorly in the secondary clarifiers, resulting is high turbidity and elevated total suspended solids leaving the clarifiers.

The graph below compares the F:M ratio using the MLVSS concentration versus the active volatile suspended solids (AVSS) value generated from an ATP analysis. This graph clearly shows that for this conventional, plug flow, activated sludge system the applied organic load exceeds design conditions by 57.5 percent. As a result, the ATP analysis immediately explains the low DO condition, odor complaints, and poor solids settling in the secondary clarifiers. There is no other analysis that could reveal these extreme operating conditions, which is why I place so much value in ATP testing, expensive as it is to conduct.

F:M Ratio Comparison Using MLVSS vs AVSS

Operators generally have no choice but to use the MLVSS number to calculate their F:M ratio. I say this because it is the only number they have available but, in fact, many industrial treatment plant operators do not have MLVSS numbers and have to use MLSS data instead to calculate their F:M ratio. Using MLSS, in place of the MLVSS concentration, further distorts the true organic loading being applied to the biological treatment system. I think, gradually, the value of ATP testing will begin to reveal itself when used by any wastewater treatment plant experiencing operating problems. Such use of ATP data would lead to a modified F:M formula as shown below.

"Modified" Food-to-Mass Ratio

F:M Ratio Using AVSS

There are two ATP kits for wastewater treatment that can be purchased from Hach as follows:

  1. QuenchGone21™ Wastewater Test Kit

  2. QuenchGone21™ Advanced Wastewater Test Kit

The only difference between the two is that the advanced kit tests for filamentous. The step-by-step instructions from LuminUltra for using either of these test kits can be downloaded here.

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