Methods to determine oxidative stability

Methods to determine oxidative stability

The auto-oxidation of fats is of paramount importance in the food industry, as the degree of lipid oxidation has remarkable consequences for the quality of the food.

The evaluation of oxidative stability faces two great difficulties: in the first place, the complexity of the reactions involved in the auto-oxidation of fats and the wide range of oxidative compounds produced cause great difficulty in their evaluation (Márquez-Ruiz et al., 2003); secondly, the oxidative stability of food determined in the laboratory cannot give an indication of the shelf life of food in practice.

The oxidation reactions consist of three phases:

  1. Initiation
  2. Propagation
  3. Termination

In the first stage free radicals are formed from unsaturated fatty acids, which combine with oxygen to form lipid peroxides. In the second the peroxides accumulate, this being the phase in which most of the unsaturated lipids are oxidized. In the last stage, the free radicals that come from the decomposition of the lipid peroxides are associated, creating non-radical compounds of low molecular mass (aldehydes, lactones, ketones, etc.) responsible for the rancid odor.

After making a quick overview of the different possibilities of evolution of auto-oxidation and the large number of products that are derived from it, we must address the problem of how to assess the development of oxidation in a fat or oil and how to express it in figures to apply a criteria of quality, acceptance or rejection, duration, average life and conservation.

Historically, methods have been created that can be divided into two groups: static and dynamic methods.

Static methods to determine oxidative stability

All these methods measure one or more functional groups that at any given moment of the oxidation process may or may not be present, and if they are the interpretation of the results is not always unequivocal. Oxidation is a dynamic process and therefore it is difficult to measure with specific data like these.

These methods provide an assessment of the punctual state of the oxidation of a fat. The most used are:

Peroxides index

It is an empirical method that assesses the oxidative capacity of a fat on iodide in acetic medium to give iodine that is titrated with bisulfate.

As described above, the oxidation process begins with the formation of hydroperoxides, which in the second phase of oxidation decompose into short chain molecules and the free radicals are coupled and form polymers. If the oxidation is not the result of a controlled acceleration, we cannot define the oxidation state of a fat from the peroxide index.


Quantifies oxidation by-products, such as high molecular weight carbonyl compounds. An oxidized and deodorized oil would be detected by this analysis.


Test of thiobarbituric acid that reacts with aldehydes such as malonic acid. A red coloration is formed that is measured spectrophotometrically. Like the previous test, it is not altered by deodorization.

Iodine index

As the oxidation of a fat progresses, the number of unsaturations decreases and therefore the Iodine index decreases.


Measures the degree of hydrolysis of fats. It is not a significant method, since there are high acidity industrial fats that do not have to be oxidized.

Absorption in the UV

In the Ultraviolet region they absorb the conjugated dienes and trienes. Natural fats do not have these structures, but instead can generate them during the oxidation process. Measurements at 232 and 270 nm allow to assess the degree of oxidation.

Absorption to IR

It has been used to detect certain chemical functions that originate in oxidative degradation.

Dynamic methods to determine oxidative stability

They are those that force the oxidation of a fat according to different procedures and measure its evolution. They accelerate a process from months to a few hours. Historically different methods have been used both to accelerate oxidation and to measure its evolution. We are going to analyze the following:

Rancimat method

Rancimat method consists of a measure of the conductivity of the volatile compounds that are formed from oxidation. The apparatus is similar to the one used in the AOM or Swift test, except that the gases that are formed are poured into a tube containing distilled water and the conductivity of the solution is measured between two platinum electrodes.

It is a standardized and commonly accepted method, but can lead to errors, some due to any trace remaining in the tubes or even the petroleum jelly used to close them, which can affect increasing conductivity. Certain free fatty acids of a low molecular weight that make up some oils can even volatilize at these temperatures (100°-120° C) also giving an increase in conductivity. On the other hand, certain antioxidants volatilize at these temperatures, which not only causes an increase in conductivity, but also are totally ineffective because they do not remain in the oil that you want to protect.

Schaal test of the stove

The Schaal method consists of subjecting a fat to temperatures of 60º-63º C. The increase in temperature acts as a catalyst accelerating the oxidation reactions, thus allowing us to measure its evolution, both organoleptically (color, smell, flavor, etc.) as per chemical analysis. Periodic index measurements are made and a graph is created showing the evolution of said index over time.

It is a very reliable method, given that by not subjecting fats to high temperatures, the evolution of oxidation is perfectly followed and the antioxidants that are volatile remain in the product and can act. One of the disadvantages is that although the use is simple, this test is very variable and is not practical as a routine analysis system.

Methods of Oxygen Absorption (RapidOxy)

The RapidOxy method consists of an accelerated oxidation process by increasing the pressure of oxygen and temperature, allowing us to determine the oxidative stability of the samples. It is carried out in oxygen pumps or special devices and the pressure drop is usually measured as a function of time. The samples are subjected to a pressure with pure oxygen of up to 700 kPa, while raising its temperature to 200° C. The temperature is maintained constant while the pressure is measured continuously until a definite drop in pressure is detected.

This method has important advantages since there is no need for expensive reagents, which are dangerous for the extraction of fats. Only a small sample volume is needed, it is also faster than other accelerated oxidation methods, saving time and money.

AOM Test or Swift Test

It is subjecting to fat at a temperature of 97.8° C in a thermostatic bath and with an air flow of 2.33 ml/sec. The fat is periodically extracted and its peroxide value is measured. The final point is the time needed to reach 100 meq/kg of IP. In the practical work reforms have been introduced to the method: there are those who establish the end point for animal fats at 20 meq/kg and maintain 100 meq/kg in vegetable oils; another variation is to express the result as an index of peroxides measured at 8 hours of test.

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