Beyond Protein: The Role of Devarda Distillation in Selective Nitrogen Determination

Nancy Beef has returned from her trip to Germany, where she helped a client perform acid hydrolysis to improve their fat determination. “This step releases encased fats that would otherwise remain inaccessible,” explains Nancy.

“Excellent work, Nancy,” says Holmes. “I wonder… Can anyone tell me about another method whereby certain compounds may remain hidden and not captured without a breakdown process?” asks Holmes. 

How can you determine organic nitrogen content?

“The Total Kjeldahl nitrogen (TKN) method has a digestion step whereby the sample is digested using sulfuric acid. The acid breaks down organic matter, converting it into ammonium, a detectable form of nitrogen that can be measured after distillation and titration. We recently compiled a selection of our best practices and top tips relating to Kjeldahl,” says Eggcule Poirot. 

“You’re on the right track, but just as hot extraction will fail to measure encased fats without hydrolysis, Kjeldahl will also be unable to detect all nitrogen compounds within a sample, even with digestion using sulphuric acid. Can you tell me what might be missed?” asks Holmes.

How can you determine inorganic nitrogen content?

After a short while, Holmes can see that the detectives are stumped. “The clue was in your description of TKN, Eggcule. You said that the acid breaks down organic matter. The compounds that are missed are inorganic matter, namely nitrates (NO₃⁻) and nitrites (NO₂⁻). These compounds are not broken down in the acidic digestion conditions used in the TKN process,” explains Holmes. 

“So, if they are not broken down by sulfuric acid, how can they be got at?” asks Cornlumbo. 

“What you need is Devarda’s alloy, which is a mixture of aluminium, copper, and zinc. This alloy was synthesized by Italian chemist Arturo Devarda, who wanted to analyze nitrate in Chile saltpeter. Deverda’s alloy can break down nitrates (NO₃⁻) and nitrites (NO₂⁻), reducing them to ammonium (NH₄⁺) in alkaline conditions,” explains Holmes.

“So, once you’ve broken down the nitrates and nitrites into ammonium, they can be quantified as part of the overall nitrogen content, giving you a better representation of the total nitrogen content, just like acid hydrolysis helps you getter a more accurate total fat content,” says Nancy Beef.

What is selective nitrogen determination?

“That’s right, but it also allows for selective reduction. Selective nitrogen determination helps you distinguish between organic nitrogen and nitrogen from nitrates and nitrites. In addition, nitrate/nitrite in samples can influence the TKN result. During digestion, nitrate can oxidize a portion of the ammonium released from the digested organic nitrogen, producing N2O and resulting in a lower recovery rate,” says Holmes.

“I didn’t realize there were so many types of nitrogen determination; I figured nitrogen was nitrogen,” says Cornlumbo.

“There are many reasons to analyze nitrogen as the different forms are essential for different applications. We have helped many of our food clients with nitrogen analysis to determine protein content. However, nitrogen is also crucial for environmental monitoring to assess pollution levels, for agriculture to optimize fertilizer use, and for wastewater treatment to ensure effluent discharge meets regulatory limits. There are specific methods for directly determining total nitrogen (TN) and ammonia nitrogen (NH3-N). TN is the combined total of total Kjeldahl nitrogen (TKN) and total oxidized nitrogen (TON). Direct determination of ammonia nitrogen enables calculation of the organic nitrogen (Norg) and total inorganic nitrogen (TIN). Luckily for us and our clients, a handy guide provides a more detailed explanation of the practical implementation of these selective nitrogen determination methods.”