van Slyke method

What is van Slyke method?

Van Slyke introduced a chemical test in 1910 to determine the amount of amino acids containing a primary amino group in protein and food hydrolysates. van Slyke method involves treating a sample in glycerol HOCH2CH(OH)CH2OH with sodium nitrite NaNO2 and acetic acid AcOH, and measuring the volume of nitrogen gas N2 generated, which is why it is commonly referred to as the van Slyke determination, van Slyke amino nitrogen determination, or the van Slyke test. Other names for this method include the van Slyke nitrous acid method, van Slyke procedure, van Slyke reaction, McLean-van Slyke method, Salkowski-van Slyke method, and van Slyke apparatus.

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van Slyke method

van Slyke method is performed for 30 minutes, during which the acidification of NaNO2 generates nitrous acid, which reacts with the primary amino group of the amino acids to form a diazotinium salt. Excess nitrous acid decomposes to water and nitrogen oxide.

The gas mixture is then passed into a modified Hempel pipet where the nitrogen oxide N2O3 is absorbed by alkaline permanganate, and the residual nitrogen molecule N2 is measured in a special gas burette.

van Slyke method can be used to determine the amount of amino acids quantitatively and the result is consistent with the Sanger method and ninhydrin. However, froth is unavoidable, so a foam inhibitor such as amyl alcohol, secondary caprylic alcohol, or phenyl ether is used.

Due to the instability of certain amino acids during the prolonged hydrolysis of proteins with 20% hydrochloric acid HCl, the amino acids determined might be inaccurate. For instance, upon boiling for 24 hours, 15 % tryptophan is inactive for this determination due to deamination, and 35.5–37.9 % of cystine nitrogen cannot be determined by phosphotungstic acid H3PW12O40. Additionally, this method is challenging for water-insoluble proteins.


Van Slyke, D. D., Proc. Soc. Experiment. Biol. Med., 1910, 7, 46