What Is the Kjeldahl Method?

The Kjeldahl method was established by the Danish chemist Kedall in 1833, and has been developed into constant, micro, and low-level Kjeldahl methods and automatic nitrogen analyzer methods, which are used to analyze the nitrogen content of organic compounds. Common methods. The theoretical basis of the Kjeldahl method is that the nitrogen content of proteins usually accounts for about 16% (12% to 19%) of its total mass.Therefore, by measuring the nitrogen content in a substance, the Total protein content (assuming that all nitrogen in the test substance comes from protein), that is: protein content = nitrogen content / 16%. ^[1]

Kjeldahl method

Protein is nitrogenous

All reagents use ammonia-free

Nitrogen Distillation Device:

as the picture shows.

1,

X = ((V1-V2) * N * 0.014) / (m * (10/100)) * F * 100%

X: the percentage of protein in the sample, g;

V1: volume of sulfuric acid or hydrochloric acid standard solution consumed by the sample, ml;

V2:

(1) The sample should be uniform. Solid samples should be thoroughly mixed beforehand, liquid samples should be shaken or stirred uniformly.

(2) When the sample is placed in the nitrogen flask, do not attach it to the neck. In case of adhesion, it can be washed down with a small amount of water to prevent the sample from being digested incompletely and the result is low.

(3) If it is not easy to present a transparent solution during digestion, cool the nitrogen fixing bottle, and slowly add 2-3ml of 30% hydrogen peroxide (H2O2) to promote oxidation.

(4) Do not use strong fire during the entire digestion process. Maintain gentle boiling, so that the firepower is concentrated on the bottom of the Kelly flask, so as to prevent the protein attached to the wall from losing nitrogen in the absence of sulfuric acid.

(5) If sulfuric acid is lacking, too much potassium sulfate will cause ammonia loss, which will form potassium hydrogen sulfate without interacting with ammonia. Therefore, when too much sulfuric acid is consumed or the fat content in the sample is too high, the amount of sulfuric acid should be increased.

(6) The effect of adding potassium sulfate is to increase the boiling point of the solution, copper sulfate is used as a catalyst, and copper sulfate is used as an indicator of alkaline reaction during distillation.

(7) The mixed indicator is green in alkaline solution, gray in neutral solution, and red in acidic solution. If bromocresol green is not available, a 0.1% methyl red ethanol solution can be used alone.

(8) Whether the ammonia is completely distilled off can be tested with a PH test paper to see if the distillate is alkaline.

(9) The absorption solution can also use 0.01 equivalent of acid to represent boric acid, and the excess acid solution is titrated with 0.01N alkali solution. When calculating, A is the number of alkali solution consumed by the reagent blank, B is the number of alkali solution consumed by the sample, and N is the alkali solution. Concentration, the rest are the same.

(10) The absorption solution using boric acid as ammonia can omit the operation of calibrating the lye, and the volume requirements of boric acid are not strict, and the use of a pipette can be eliminated, and the operation is relatively simple.

(11) When adding concentrated alkali to the distillation flask, brown precipitates often appear. This is because the decomposition-promoting alkali reacts with the added copper sulfate to form copper hydroxide, which is decomposed to form a precipitate of copper oxide after heating. Sometimes copper ions interact with ammonia to form a dark blue conjugate [Cu (NH3) 4] 2+

(12) The essence of this calculation method is to measure the nitrogen content and then make an estimation of the protein content. This method can only be used to estimate protein content when the composition of the test substance is protein.