Kjeldahl analysis for the determination of nitrogen and protein

The nitrogen/protein analysis according to Kjeldahl is up to date still the reference method in analytics. With its high degree of versatility, the method is suitable for many applications, e.g. food analysis, feed analysis, soil or water analysis. In addition to the versatility, the very high precision also speaks for the method. Particularly in case of highly inhomogeneous sample material, there is hardly any alternative to Kjeldahl due to the high weight of the sample. The Kjeldahl process was developed as early as 1883 by Johan Kjeldahl and is still used in this form today. It can be separated into 3 major process steps:

  •     Digestion of the samples with sulphuric acid
  •     Distillation of the digestion solution with steam
  •     Titration of the distillate and calculation of the result

In the classic application, manual laboratory heaters are used, as well as round bottom flasks for digestion and Erlenmeyer flasks for distillation. With the complete automation of the method using the digestion block and steam distillation system with integrated titration, the sample throughput can be increased significantly. At the same time, the repeatability of the analysis is increased and the entire work process is much safer for the laboratory staff. The diagram below shows how Kjeldahl analysis with automatic analysers works step by step.

Automation of the reference method for nitrogen and protein analysis

Step 1 - Sample weighing

Weighing in sample on N free paper

Step 2 - Sample transfer into digestion tube

Transfer of sample with weighing boat into digestion tube

Step 3 - Digestion

Addition of salt to raise boiling point and as catalyst, e.g. KJELCAT Cu (K2SO4 + CuSO4)

Step 4 - Digestion

Addition of sulphuric acid

Step 5 - Digestion

Digestion at boiling point, for about 60 to 180 minutes

CnHmNx + H2SO4 → n CO2 + ½ m H2O + ½ x (NH4)2SO4 (solv)


Step 6 - Distillation

Dilution of sample solution with water to prevent strong reactions by following addition of caustic soda.

Step 7 - Distillation

Addition of caustic soda to release ammonia, today this is done automatically in modern steam distillation units like VAPODEST

NH4+ + OH-NH3↑ + H2O

Step 8 - Distillation

Seperation of ammonia by steam distillation and trapping the condensated ammonia-water solution in boric acid

NH3 + H3BO3NH4+ + H2BO3-

Step 9 - Titration

Quantitative determination of nitrogen by titration with sulfuric acid or hydrochloric acid. With direct pH measurement with pH electrode or with pH indicator.

NH4+ + H2BO3- + HCl → NH4CL + H3BO3

Step 10 - Calculation

Calculation of nitrogen content:

% N = (1,4007 ∗ ceq ∗ (V - Vb)) / E

ceq - H+ Ion concentration of standard volumetric solution: hydrochloric acid c = 0,1 mol/l

alternativ: sulfuric acid ceq = 0,1 mol/l

V - Consumption volumetric standard solutions sample (ml)

Vb - Consumption volumetric standard solution blank (ml)

E - Weight (g)


Calculation of protein content:

% Crude protein = % N * PF

Examples of protein factors (PF):

6,38Milk, cheese, milk powder products, milk products
6,25Meat, fish, poultry, egg, vegetable, fruits, different types of grain, corn, legumes, feed
5,71Soy beans
5,7Wheat and wheat flour
5,4Oilseeds, nuts


Note: The listed protein factors are often applied. Depending on regulation the analysis is based on, different protein factors must get applied. Especially in grain, nuts and oilseeds or single components of the above described sample types.


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