Oceanography

4. Procedure

25 ml of filtered sample is taken in a conical flask. To this solution 0.5ml of sulphanilamide solution is added and the solution is stirred well. After not less than 3 min and not longer than 8 min 0.5ml of diamine solution is added and the content of the conical flask is thoroughly mixed. After 10 minutes, the OD of the red azodye developed is measured in spectrophotometer at 543nm. The colour is stable for 2 hrs. The values of OD recorded for triplicate samples are registered in the data book.

Formation of NO₂-N working solution

0.345 g of anhydrous NaNO₂ is dissolved in distilled water and the volume is made up to 100ml. 1ml of chloroform is added as preservative. 1 ml of this solution contains 70µg or 5 µg at NO₂-N. The NO₂-N working solution is prepared from the stock solution by diluting 1ml of solution to 100ml. 1ml of this solution contains 0.7µg or 0.05µg-at N.

Formation of calibration curve

The working solution is further diluted with distilled water to get the following standard solutions.

i. 0.1ml working solution made up

To 100 ml with distilled water = 0.05µg-at NO₂-N/l

ii. 0.2 ml of working solution made up

To 100ml with distilled water =0.1µg-at NO₂-N/l

iii. 0.5 ml of working solution made up

To 100 ml with distilled water =0.25µg- atNO₂-N/l

iv. 1 ml of working solution made up

To 100ml with distilled water =0.5µg-at NO₂-N/l

v. 2ml of working solution made up

To 100ml with distilled water = 1µg-at NO₂-N/l

25ml of the above diluted working solution is treated with sulphanilamide and diamine solution to get an azo dye. The values of OD recorded for the different concentrations are tabulated. The OD is taken on the ‘X’ axis and concentration is plotted on the ‘Y’ axis. From the linear graph drawn between OD and concentration of NO₂-N, it is possible to assess the NO₂-N content of the given seawater sample.