Nitrite-API colorimetric assay

Nitrite is measured with the standard Greiss test. In this test, sulfanilamide reagent is converted to a diazonium salt by nitrite (NO2) in the water sample. The diazonium salt is then reacted with NED (N-1-napthylethylene diamine dihydrochloride) reagent to form an azo dye which has a purple/magenta color that is measured at 520-550 nm (suitable for the colorimeter 528 nm green LED).

Absorbance spectrum

• Reported values in literature around 540nm
• Measured absorbance peak at 530nm
• Open Colorimeter measurement LED:  520nm

Materials

• Open Colorimeter with 520nm LED board
• API nitrite test kit
• 1,000 ppm N (3,280 ppm as NO₂) standard, Labchem Cat # LC17930-7
• Distilled water
• Cuvettes
• 6 x 5 mL capped tubes
• 2 x 100 mL volumetric flask
• 1 mL adjustable pipette

Prepare nitrite calibration solutions: 0-2ppm

• Prepare 100 ppm NO₂: 3.05 mL of 3,280 ppm standard in 100 mL volumetric flask. Fill to line with distilled water.
• Prepare 2.0 ppm NO₂: 2.0 mL of the 100 ppm working stock in 100 mL distilled water in volumetric flask. Fill to the line with distilled water.
• Dilute the 2 ppm working stock as outlined below

Method

• To each 5 mL sample, add 5 drops of API Nitrite Test Solution
• Cap the tube and invert several times to mix.
• Wait 5-10 mins for the color to fully develop.
• Make sure the Open Colorimeter has the 520nm LED installed
• Turn on the Open Colorimeter and select absorbance. Blank the colorimeter with the 0 ppm nitrite solution and measure absorbance of all of the solutions.

Calibration data

{
  "Nitrite": {
    "units": "ppm",
    "led": "520",
    "fit_type": "polynomial",
    "fit_coef": [0.1311, 1.2591, 0.0],
    "range": {"min": 0.0, "max": 1.4}
  }
}