Monitoring Nitrite Levels in Engine Coolant

Engine coolant is a critical fluid that is essential for the optimal performance of engines in motor vehicles. Several studies have reported that, behind lubrication failure, a major cause of engine damage is related to engine coolant.1

Formulations of engine coolant (also known as antifreeze) consist of glycol, water, and several additives.2 Although they differ in formulation and chemistry, all the additives in an engine coolant are used to prevent corrosion and promote operational efficiency. There are several testing procedures available for coolant analysis.

A common additive in coolant formulations is nitrite (NO2.) This additive prevents pitting from occurring, creating a protective layer of oxide patina on cylinder liners.1 However, nitrite levels must be frequently tested to maintain sufficient levels as this additive is consumed during use.

Several techniques are commonly employed to test nitrite levels in engine coolant, including ion chromatography and the paper test strips method. The paper test strip method is a relatively straightforward technique that can be used to provide quick manual analysis of nitrite levels.

Ion chromatography provides quantitative analysis of nitrites and other anions in engine coolants.3 Another technique that can provide an accurate determination of nitrite levels in coolant is the UV/Vis technique, which offers benefits such as accuracy, speed, and automation, improving high-volume testing performance.

There are several methods cited in the literature for the analysis of nitrites. In this article, EPA method 354.1 was used as a template to accurately determine nitrite levels as ppm NO2 - in glycol-based aqueous engine coolant using the PerkinElmer LAMBDA® 365+ UV/Vis Spectrometer (Figure 1). All data was processed with UV Winlab software.

PerkinElmer LAMBDA 365+ Spectrometer.

Figure 1: PerkinElmer LAMBDA 365+ Spectrometer. . Image Credit: PerkinElmer


According to EPA 354.1, nitrite levels in an aqueous solution can be determined by a diazotization reaction with sulfanilamide under acidic conditions.4

The product of this reaction is then coupled with N-(1-Naphthyl)ethylenediamine dihydrochloride to obtain a purple-colored product that can be detected spectrophotometrically at 540 nm.

This method reveals nitrite concentration as it has a directly proportional relationship to the resulting UV/Vis absorbance at 540 nm based on the principle of Beer’s Law. Reagents used for the preparation for samples and standards according to EPA 354.1 are listed below:

  1. 1.N-(1-Naphthyl)ethylenediamine Dihydrochloride
  2. Anhydrous Sodium Nitrite
  3. Sulfanilamide
  4. Nitrate and nitrate-free distilled water
  5. Concentrated Hydrochloric Acid
  6. Chloroform
  7. Sodium Acetate

EPA 354.1 reports nitrites as mg/L NO2 - N (Nitrite - Nitrogen) over a calibration range of 0 – 10 mg/L in water, wastewater, and sludge. Reagents and standards were used to report nitrite levels as ppm NO2 in this article.

Ready-made reagents and standards are available from numerous chemical suppliers, and these were used where possible.

As the nitrite concentration in engine coolants exceeds 1000 ppm in a new sample, a dilution factor will need to be applied for the analysis.

The team determined that a UV/Vis cuvette with a nominal path length of 1 mm should be employed for this application rather than the standard 10 mm cuvette used for the EPA method. This approach confers the benefits of analyzing more concentrated samples and reducing the total dilution factor.


A buffer color change reagent was prepared by adding 0.25 g N-(1-Napthyl) ethylenediamine Dihydrochloride to 25 mL sulfanilamide reagent (1 % (w/v) sulfanilamide in 10 % HCl).

This reagent is light-sensitive and should be stored appropriately. The reagent was tested for stability and had a shelf life of up to two weeks.

A premade 1000 ppm nitrite standard was purchased to make the calibration standards. Nitrite and nitrate-free water were used to dilute the premade nitrite standard. A working standard of 100 ppm NO2 was produced. Table 1 shows the range of dilutions used in the application.

Table 1. Calibration range for nitrite standards. Source: PerkinElmer

Description NO2- concentration (ppm)
Standard 1 0.25
Standard 2 0.50
Standard 3 1.00
Standard 4 5.00
Standard 5 10.00


For sample preparation, 5 mL of nitrite and nitrate-free distilled water was added to a 10 mL volumetric flask. To this volume of water, 40 µL of sample of standard was added, followed by 250 µL of the buffer color change reagent.

Distilled water was then added up to the mark on the flask, and a 250-fold dilution factor was achieved after shaking.

Samples and standards were allowed to sit for thirty minutes before analysis. This allowed the color to develop and stabilize after buffer color reagent addition. A Wavelength Quant method on UV Winlab software was created using a wavelength of 540 nm.

Standards and samples were measured in a UV/Vis cell with a nominal pathlength of 1 mm.


The spectra of the nitrite standards as well as the resulting calibration curve, are shown in Figures 2 and 3. The linear regression coefficient (R2) obtained from the calibration curve displayed in Figure 3 is 0.9996, indicating an excellent level of data correlation within the specified calibration range.

Overlay of nitrite standard spectra (700 nm – 400 nm).

Figure 2: Overlay of nitrite standard spectra (700 nm – 400 nm). Image Credit: PerkinElmer

Nitrite calibration curve in ppm.

Figure 3: Nitrite calibration curve in ppm. Image Credit: PerkinElmer 

Table 2  shows the values calculated for various coolant samples using the UV/Vis calibration method. The values were compared to reference samples with a known nitrite value to determine the accuracy of the method. 

The 250-fold dilution factor was applied to the results to calculate the final adjusted amount. A 1000 ppm control sample was included in the middle of a sample set as well as at the end of the sample set to monitor the accuracy of the analysis.

Sample values agreed well with the reference values after applying the 250-fold dilution factor.

Table 2. Sample result set. Source: PerkinElmer

Sample Adjusted Nitrite (ppm) Reference Value (ppm)
Coolant 1 205 193
Coolant 2 1180 1225
Coolant 3 628 620
Coolant 4 1669 1677
Coolant 5 1404 1388
1000 ppm std (QC) 1023 1000



High accuracy quantitation of nitrite levels in engine coolant was achieved in the application by adapting EPA method 354.1 using PerkinElmer’s equipment. By using ready-made reagents, sample and standard preparations were simplified.

Automating preparation procedures and improving the efficiency and accuracy of studies is made possible by using fewer reagents. This has important implications for future work and for enhancing the efficiency of current analytical methods.

In conclusion, the coolant additives did not appear to significantly affect results accuracy when compared to real-world samples. This demonstrates that the method discussed in this article is suitable for the accurate determination of nitrite levels in the engine coolant sample matrix.

PerkinElmer’s equipment is useful for additive analysis. UV WinLab and LAMBDA 365+ software also provides a reliable solution for the accurate determination of nitrite levels in engine coolant. 


  1. Beal, RE., (1993) Engine Coolant Testing: Third Volume, American Society for Testing and Materials, Philadelphia, PA.
  2. ASTM Standard D6210, (2017) “Standard Specification for Fully-Formulated Glycol Base Engine Coolant for Heavy-Duty Engines,” ASTM International, West Conshohocken, PA, DOI: 10.1520/D6210-17,
  3. ASTM D5827, 2015, (2015) “Standard Test Method for Analysis of Engine Coolant for Chloride and Other Anions by Ion Chromatography,” ASTM International, West Conshohocken, PA, DOI: 10.5120/D5827-09R15,
  4. Methods for the Chemical Analysis of Water and Wastes (MCAWW) (EPA/600/4-79/020), EPA 354.

This information has been sourced, reviewed and adapted from materials provided by PerkinElmer.

For more information on this source, please visit PerkinElmer.


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