Wet Chemistry vs Express NIR Amino Acid Analysis: Making the Informed Choice

Amino acid analysis is an important part of any technologist's work. There are two kinds — wet-chemistry and express analysis. They differ substantially from one another, and this is important to bear in mind when choosing an analysis of feeds, premixes, PVMS, amino acid substances, and meat, fish or other products that are not a single individual plant.

The quality of the result depends on which kind of analysis you choose. Wet-chemistry has a smaller error and depends on the standard of the laboratory carrying out the work. The quality of express analysis, by contrast, is affected by a great many factors. Moreover, it can only be made reasonably reliable for single-component plant products.

Let us examine each kind of analysis in more detail.

Wet-chemistry analysis

Wet-chemistry analysis is based on a particular chemical property of the substance:● on an interaction with a reagent such that the product formed can be analysed;● or on the ability of indicators to react to the substance;● or on the action of solvents on the substance being analysed.

Amino acids specifically react with substances known as derivatising agents. These reagents convert the amino acid into a form that is easy to analyse with instruments.

An important stage in wet-chemistry analysis is sample preparation. During this stage, a liquid containing amino acids is obtained from a feed, food, solid or other sample. On average, this process takes 1–3 days.

The following equipment is used to carry out wet-chemistry analysis:● a chromatograph (a chromatograph made solely for amino acid analysis is sometimes called an amino acid analyser),● capillary electrophoresis (this is also chromatographic equipment),● a spectrophotometer.

Since the method is based on a direct interaction with a reagent and each amino acid is analysed separately, wet-chemistry analysis is quite accurate (a relative error of 3–5%).

The wet-chemistry method can determine amino acids in any product.

Express analysis

The express method is based on the spectral characteristics of the individual components that make up a plant. Such components include protein, fat, fibre, moisture and so on.

During such an analysis, the functional groups of organic substances, under infrared light, produce specific maxima and minima in the spectrum (the spectrum being the instrument's graphical response to a change in wavelength).

Specialists have learnt to find a certain relationship between the absorbance values (the Y axis) at these maxima and the content of the substance sought (say, protein in soybean cake).

To build such a relationship, the instrument (the express analyser) must be supplied with at least 100 protein values for soybean cake obtained by wet-chemistry methods (and, for a good result, 1,000 values). We shall henceforth refer to this relationship as a calibration.

Using specialised software, the instrument derives this relationship. When an unknown sample is analysed, its spectrum is recorded and the required value can be calculated. All these results are computed mathematically and can only be made more accurate with a large number of known samples. On the whole, the error of such analyses is quite large (5–30% of the wet-chemistry value).

Express analysers come in various kinds. Infrared express analysers are most often designated by the abbreviation NIRS (near-infrared spectroscopy). They are produced by many companies (FOSS, Perten, InfraTec, Infrascan and others).

The accuracy of express analysis depends directly on the quality of the instrument's calibration. And here is why.

It is important to calibrate an express analyser separately for each type of product (separately for wheat, for soybeans, for soybean cake, for soybean meal, and so on). This is because they differ in their content of protein, fat, fibre and organic acids — and it is precisely these that determine the spectrum recorded by the IR analyser.

Calibrations for protein, fibre, fat and moisture are fairly common, whereas calibrations for amino acids are very rarely carried out in Ukraine.

Often the firms carrying out the analysis install the express analyser in Ukraine, record the spectrum, and send it to partners abroad. There it is fed into the calculation system, and the result is sent back to Ukraine.

Another important factor is who performed the calibration. Only someone who has both an IR analyser and wet-chemistry equipment, and who can afford the costs involved in such a procedure, can produce high-quality calibrations. At present I know of only one manufacturer in Ukraine that independently calibrates its own equipment for determining protein and moisture. To do so, the enterprise collects samples on which it performs wet-chemistry analyses over the course of a year.

The vast majority of calibrations are installed by the supplier. It is practically impossible to establish how often the user updates them. Manufacturer support for updating calibrations is expensive (from €20,000 a year). This therefore leaves scope for "cost-cutting", which has a detrimental effect on the result.

Importantly, IR analysers analyse only plant-based raw materials. If a product contains components from various crop raw materials (feeds, PVMS, premixes and so on), an IR analyser is generally not used. The same applies to synthetic amino acids.

So, to summarise:

How to tell which method was used for the analysis

1. Ask whoever performed the analysis.

2. If results for the total amino acid content in a feed or crop raw material are provided within two days, there is a strong likelihood that it is an express analysis. Wet-chemistry analysis takes a minimum of two full days, and for some amino acids three or four — and that is provided the sample was submitted for analysis on the same day it was received.

3. By the test report:● The laboratory must state the method of analysis in the document (this is a requirement of the ISO 17025 standard). The simplest way to check it is to look it up online. Search engines will show you the full name or description of the standard, which usually specifies the method used.● Sometimes the report specifies the equipment on which the result was obtained. If NIRS is indicated, it is an express analysis. If chromatographic equipment is indicated, it is a wet-chemistry analysis.

4. Ask for the chromatogram of the analysis. This can only be obtained with the wet-chemistry method.

Which analysis to choose

If you are analysing a multi-component plant or a product of non-plant origin, then choose wet-chemistry analysis.

If you are analysing a specific plant (wheat, maize, soybean meal, soybeans, sunflower cake, and so on), you may use express analysis. But be sure to check that the equipment on which the analysis will be carried out has been properly calibrated.

Our laboratory carries out amino acid analysis by the wet-chemistry method in any product.

We look forward to working with you!

Anastasia Shuparska, Head of the Halomlab Laboratory

15 April 2021