Kjeldahl Method for Determining Nitrogen
Digestion Titration The Kjeldahl method was developed over 100 years ago for determining the nitrogen contents in organic and inorganic substances. Although the technique and apparatus have been modified over the years, the basic principles introduced by Johan Kjeldahl still endure today.
Kjeldahl nitrogen determinations are performed on a variety of substances such as meat, feed, grain, waste water, soil, and many other samples. Various scientific associations approve and have refined the Kjeldahl method, including the AOAC International (formerly the Association of Official Analytical Chemists), Association of American Cereal Chemists, American Oil Chemists Society, Environmental Protection Agency, International Standards Organization, and United States Department of Agriculture.
The Kjeldahl method may be broken down into three main steps: digestion, distillation, and titration.
Digestion is accomplished by boiling a homogeneous sample in concentrated sulfuric acid. The end result is an ammonium sulfate solution. The general equation for the digestion of an organic sample is shown below:
| Organic N + H2SO4 → |
| (NH4)SO4 + H2O + CO4 + other sample matrix byproducts |
Distillation: Excess base is added to the digestion product to convert NH4 to NH3 as indicated in the following equation. The NH3 is recovered by distilling the reaction product.
ammonium
sulfate | heat | ammonia
gas |
| (NH4)2SO4 + 2NaOH | → | 2NH3 + Na2SO4 + 2H2O |
Titration quantifies the amount of ammonia in the receiving solution. The amount of nitrogen in a sample can be calculated from the quantified amount of ammonia ion in the receiving solution.
There are two types of titrationback titration and direct titration. Both methods indicate the ammonia present in the distillate with a color change.
In back titration (commonly used in macro Kjeldahl), the ammonia is captured by a carefully measured excess of a standardized acid solution in the receiving flask. The excess of acid in the receiving solution keeps the pH low, and the indicator does not change until the solution is "back titrated" with base.
| ammonia | standard
sulfuric acid
acid | | excess
ammonium
sulfate | sulfuric
acid |
| 2NH3 + | 2H2SO4 | → | (NH4)2SO4 + | H2SO4 |
| (no color change) |
ammonia
sulfate | measured
excess
acid | measured
sodium
hydroxide | | ammonium
sulfate |
| (NH4)2SO4 + | H2SO4 + | 2NaOH | → | Na2SO4 + (NH4)2SO4 + 2H2O |
(color change occurs) |
In direct titration, if boric acid is used as the receiving solution instead of a standardized mineral acid, the chemical reaction is:
ammonia
gas | boric
acid | | ammonium-
borate complex | excess
boric acid |
| NH3 + | H3BO3 | → | NH4 + H2BO-3 + | H3BO3 |
| (color change occurs) |
The boric acid captures the ammonia gas, forming an ammonium-borate complex. As the ammonia collects, the color of the receiving solutions changes.
ammonium-
borate
complex | | sulfuric
acid | | ammonium
sulfate | | boric
acid |
| 2NH4 | + | H2BO-3 | + | H2SO4 (NH4)2SO4 | + | 2H3BO3 |
| (color change occurs in reverse) |
The boric acid method has the advantages that only one standard solution is necessary for the determination and that the solution has a long shelf life.
Kjeldahl System for Protein Analysis
Distilling Apparatus for Kjeldahl Determinations
