Kjeldahl Method

The Kjeldahl Method is used to determine the amount of Nitrogen in organic materials. The nitrogen is used as an indicator of protein since fats, dietary fibers and carbohydrates do not contain nitrogen.


There are three steps in the Kjeldahl method: digestion, distillation and titration.


In the digestion step, the organic material is combined with sulfuric acid under high heat. The nitrogen is converted into ammonium sulfate. Catalysts may be use to speed up the reaction or to raise the boiling point of the sulfuric acid.


The distillation process involves removing the ammonium ions from the solution in the previous step by conversion of the ammonium gas, through heat and acid neutralization, into a second solution of hydrochloric acid.


The final step is the combining of the hydrochloric acid solution from the previous step with sodium hydroxide. The amount of Nitrogen is calculated by subtracting the amount of Base used to neutralize the known amount of Acid. The difference is the amount of nitrogen contained in the solution.

Protein Determination

This method is an international standard for determining protein in organic material that does not have nitrogen or nitrogen containing compounds. Protein is the typically the only source of nitrogen since nitrogen is not contained in dietary fibers, such as Chiten and Cellulose among others, fats and carbohydrates.

The standard calculation to determine the percentage of protein is to multiply the percent of nitrogen by 6.25, or 1/0.16. Protein consists, on average, of 16% Nitrogen. Higher accuracy may be obtained by adjusting this protein factor according to amino acid composition.


Fischer, Robert B. Quantitative Chemical Analysis. 2nd ed. Philadelphia: Saunders, 1961: 229-31.

Last Edited July 10, 2007