Accelerated oxidation tests – the RapidOxy method


The oxidation process that affects the compounds present in foods for animal nutrition alter their qualities, both organoleptic and nutritional, producing a decrease in the useful life of the final product.

The ingredients most sensitive to oxidation are those that contain double bonds, such as fats, oils and other essential nutrients (fatty acids, vitamins, carotenoids, etc.).

The oxidation and consequent formation of free radicals in the oxidation process cause a reduction of the nutritive value of the feed and its ingredients, but also produces a decrease in the immunity of the animals and, consequently, augments the appearance of diseases and increases in mortality.

A high intake of feed with a high content of oxidized lipids and polyunsaturated fatty acids, as well as pro-oxidant components, contribute to the in vivo oxidation of animals and the post-mortem oxidation of meat (Morrissey et al., 1998 or Stem et al., 2008).

This is very important, since lipid oxidation and the consequent oxidative rancidity are one of the main causes of food spoilage forhuman consumption, along with microbial development.

Therefore, the methods that determine the oxidative stability (resistance of a sample to oxidation) are very important, particularly when evaluating the effectiveness of antioxidants to delay the oxidation of these products in animal feed, these being used to investigate the useful life of the product and also in the assurance and quality control during the development and subsequent launch of the product.

In this post, we work with several accelerated oxidation methods (Rancimat method, Schaal method) of which we have already discussed in previous articles, however today we are going to focus on another method of accelerated oxidation: the RapidOxy method.

RapidOxy is an accelerated oxidation test that corresponds to the methods of oxygen absorption, which in recent years has become more relevant, especially in the animal nutrition industry, although it is also used in other industries such as food or cosmetics .

RapiOxy is based on the increase of oxygen pressure and temperature inside a sealed chamber, allowing us to determine the oxidative stability of the samples studied. It is carried out in oxygen pumps or special devices and the decrease in oxygen pressure is generally measured as a function of the time elapsed. The samples are subjected to a certain pressure with oxygen while their temperature rises, usually up to 200º C. The temperature remains constant, while the pressure is measured continuously until a definite drop in pressure is detected. Oxidation stability characterizes the resistance of the sample against oxygen.

With this accelerated oxidation method, only a small sample is needed to get accurate results in short periods of time, and without having to prepare the samples in advance. It is a very useful procedure for investigations on useful life, quality control of the raw materials and during the development of the products.

RapidOxy accelerated oxidation method procedure

  • The sample is deposited in a small sealed test chamber, where it is subjected to a pressure with pure oxygen of up to 700 kPa, while raising its temperature to 200 ° C.
  • The temperature is kept constant and the pressure is measured continuously until a defined pressure drop is detected.
  • The result is indicated as the induction period (IP) which is the time elapsed between the start of the test (when the heating of the container where the sample begins) and the point of rupture (instant where the oxidation increases rapidly, which causes a rapid oxygen consumption and therefore a defined pressure drop), which informs us of the oxidation stability of the sample tested.

Advantages of RapidOxy accelerated oxidation method

RapidOxy accelerated oxidation method has important advantages over other alternative methods:

  • It is not necessary to prepare samples and, therefore, there is no need for expensive or dangerous reagents for the extraction of fats.
  • We can work with liquid, solid and semi-solid samples. Normally, only 5 mL of sample is needed, that is, a small sample volume.
  • It is faster than other methods of accelerated oxidation, so it saves time and money.
  • Cleaning is quick and easy.
  • Your safety has been approved by the German Federal Institute for Materials Research and Testing.
  • It includes  free software that is supplied with the equipment, which allows the transfer of the results to the computer, obtaining graphs in real time which allows a quick comparison of the results.

Main uses of the RapidOxy method

  • Evaluate oxidative stability at high temperatures.
  • Predict the useful life of the product.
  • Control accelerated oxidation.
  • Detect the presence of antioxidants.
  • Control the conditions of packaging and storage.

RapidOxy applications in the animal nutrition industry:

  • Determine oxidative stability of vegetable oils and animal fats (eg. oils, lard, tallow, fish fats, etc.).
  • Determine oxidative stability of solid products (eg. compound feeds, calcium soaps, meat meals, fish meal, etc.).

Oxidation of fats affects both the feed with which the animals are fed, and the animal’s own organism and / or subsequently the quality of the products of animal origin for human consumption.

RapidOxy accelerated oxidation method is a type of assay that becomes increasingly relevant due to its multiple advantages and that can be used for a wide range of products in the animal nutrition industry.

Classification of food antioxidants: secondary antioxidants

Continuing from the previously discussed topic of primary food antioxidants, we will talk about secondary antioxidants.

For some context, we will say that food antioxidants  act by exerting an inhibitory action on peroxidation, transforming free radicals into more stable compounds.

In this case, secondary antioxidants delay oxidation by chelating metals, regenerating primary antioxidants, decomposing hydroperoxides or eliminating oxygen (Johnson 1971, Labuza 1971 and Gordon 1991).

Secondary antioxidants can be classified into the following groups:

A) Antioxidant oxygen receptors

Ascorbic acid and ascorbates

E-300 Ascorbic acid

E-301 Ascorbate sodium ascorbic acid sodium salt (L +)

E-302 Calcium ascorbate Calcium salt of ascorbic acid (L +)

E-303 Ascorbyl Diacetate

The L-ascorbic or vitamin C, is a white solid which is odorless, highly soluble in water and insoluble in fats and oils. It can act as an oxygen receptor, although its form of action depends on the concentration and the product in which it is used. Accordingly, ascorbic acid can be used:

  1. For chelating, when there is low water activity.
  2. As a receiver or oxygen eliminator of the medium. In the presence of oxygen and metal ions, in an aqueous medium, it is oxidized to dehydroascorbic acid, being more effective at low oxygen levels.
  3. As asynergistic of type l antioxidants.
  4. As an agent that helps the formation of radicals and act, therefore, as prooxidant.

The food industry uses ascorbic acid for the production of a wide variety of food products, including canned or bottled products, beverages, vegetable oils, fruits, butter, vegetables, cured meat and canned fish.

Erythorbic acid (isoascorbic acid)

E-315 Erythorbic acid (isoascorbic)

E-316 Sodium erythorbate (sodium isoascorbate)

Erythorbic acid is the D- isomer of ascorbic acid. It has no vitamin activity and is found naturally in food. Erythorbic acid as well as its sodium salt are used in the stabilization of nitrates and nitrites in cured meat products, dehydrated fruits and vegetables, and as synergists of tocopherols in fats and oils (Nakao, et al., 1972, Movaghar, 1990) .

Erythorbic acid, in combination with citric acid, can be used, as an alternative to sulphites in frozen seaoods, vegetable salads and apples.

B) Chelating agents

In this group we include substances that have antioxidant action through a specific mechanism, which consists of the sequestration of traces of metals present in food. The chelating agents have the mission of capturing the metal ions, forming complexes that remain soluble and innocuous, which is of great importance in the food industry, avoiding undesired effects in the production processes or in the final product.

In the process of selecting a chelating agent, in addition to taking into account toxicological and sensory aspects (color and taste), other aspects of interest must be assessed, such as the characteristics of the medium (pH) since they significantly influence the effectiveness of the chelation and solubility.


E-338 Phosphoric acid

E-339 Orthophosphoric salts

E-340 potassium dihydrogen phosphate

E-341 Calcium orthophosphate

E-341iii) Tricalcium Orthophosphate

Phosphoric acid and its salts are used in the food industry as chelating agents, as stabilizers of emulsions and as anti-hardening agents.

The excretion of phosphates takes place, especially in faeces, in the form of calcium phosphate. Because of this, an excessive intake of phosphates can produce  bone mass and decrease in calcium.

Tartaric acid

E-334 Tartaric acid

E-335 Sodium tartrate

E-336 Potassium Tartrate

E-337 Mixed Tartrate of Potassium and Sodium / Salt of Seignett

Tartaric acid is found naturally in many fruits, and is also a byproduct of winemaking. Tartaric acid is absorbed almost completely in the intestinal tract, being metabolized significantly, in body tissues, giving rise to CO2.

Citric acid

E-330 Citric acid

E-331 Sodium Citrate / Sodium Citrate

E-332 potassium citrate

E-333 Calcium citrate

Citric acid and its salts are used as chelating agents. They are used as synergists with primary antioxidants and with oxygen receptors at levels of 0.1-0.3%. In fats and oils, citric acid forms chelates with metal ions at concentrations of 0.005-0.2% (DziezaK, 1986).


E-322 Lecithin-N

Lecithin (phosphatidyl choline) is a phospholipid found in natural products, with a percentage of 1-2% in many vegetable oils and animal fats. The main source is soy. Commercial lecithin is formed by a mixture of phospholipids.

Lecithin acts as a powerful synergist in fats and oils, along with primary antioxidants and oxygen receptors at temperatures above 80 ° C. When there are low concentrations of antioxidants, lecithin is more effective. It is also very effective when forming ternary mixtures with vitamins E and C, to such an extent that the induction times in the oils increased about 25 times when adding 500 ppm of vitamin E and 1000 ppm of vitamin C (Loliger, 1991). Similar effects are found in mixtures containing ascorbyl palmitate, lecithin and α-tocopherol (Hudson and Ghavami, 1984).

C) Eventual antioxidants

Amino acids

Amino acids are effective both as primary antioxidants and as synergists (Bishov and Henick, 1975). The antioxidant activity of many amino acids is dependent on concentration and pH. At high concentrations and low pH they act as pro-oxidants, while at low concentrations and high pH they have antioxidant properties.

Methionine, histidine, proline, tryptophan, glycine and lysine are effective in fats and oils.

Spice extracts

Spice extracts are a potential source of natural antioxidants. They are effective in fat, meat and bakery products. Rosemary and sage bring good antioxidant properties to lard. Eugenol is the main component of clove and curcumin, the main pigment in turmeric, all three  of which have antioxidant properties (Cort, 1974b). Spice extracts have a strong smell, color and flavor, so they can only be used in foods that are compatible with these characteristics.

Vitamin A

Vitamin A has a very limited use as an antioxidant due to its tendency to oxidize when exposed to light or air, conditions under which the vitamin becomes pro-oxidant.

Retinol is a form of vitamin A. It belongs to the group of retinoids and is widely used for its high effectiveness in fats and oils when stored in the dark. In addition, this substance inhibits the formation of free acids in vegetable oils. Retinol is found in all animal tissues, mainly in the liver, as well as in eggs and milk. The liver is the primary storage site for vitamin A. The recommended daily intake is 750 mg / kg-pc (FAO / WHO, 1967).

Incorporation of antioxidants

One of the main problems that arise when using antioxidants is the achievement of an effective and complete distribution in the food, in such a way that they can come into contact with the lipids and act effectively. This problem worsens when the antioxidant has to be added in foods with a low lipid content and with a defined structure.

The addition of antioxidants is easier in more or less viscous products (oils and fats), or have physical structures that allow homogenization (chopped products, emulsions, etc.).

Classification of food antioxidants: Primary antioxidants

As we have already commented on in other occasions, antioxidants are any substance capable of inhibiting, delayingor preventing the development of rancidity in food or other deterioration of aromas due to oxidation. According to this definition, antioxidants do not improve the quality of food, but their use simply aims to maintain food quality.

To inhibit, reduce or delay the oxidation of lipids, it is necessary to act against one or more of the factors that favor their development.

In a broad sense, according to the above definition, an antioxidant is considered to be any substance or action procedure that helps to limit the speed and / or extension of oxidative processes, so it can be considered as such, not only the chemical compounds that can be added to the product but also: vacuum packaging, in an inert gas atmosphere or even freezing.

According to what has been said, three types of antioxidants could be considered, according to their mechanism of action. Two of them are associated with the addition of chemical compounds that are, in addition, those that we will consider in this article. The third type of antioxidant owes its action to modifications of certain factors in the food and / or its processing and will not be analyzed here by us.

Classes of antioxidants

For its mechanism of action  two main types of antioxidants (I and II) can be considered The primary (type I) are those that break the chain reaction of oxidation through the donation of hydrogen and the generation of more stable radicals. In contrast, secondary antioxidants (type II) are those that delay oxidation through other mechanisms, such as metal chelation, the regeneration of primary antioxidants, the decomposition of hydroperoxides and the elimination of oxygen, among others. This mechanism of antioxidant activity has been studied by numerous researchers (Johnson, 1971, Labuza, 1971 and Gordon, 1990).

Primary antioxidants or type I

These antioxidants are those which break the chain reaction of oxidation through the donation of hydrogen and the generation of more stable radicals.

The addition of these compounds to food should, by itself, imply an increase in the induction period.

This increase is directly related to the amount of antioxidant added up to a certain concentration, since, sometimes with higher proportions, an opposite effect is achieved.

The effectiveness of the activity of these antioxidants depends both on the antioxidant itself and on the medium in which it acts. Thus, it has been proven that in phenolic antioxidants its activity is favored when vacuum packaging is done, since the level of available oxygen is very low. However, this offer little protection when the concentration of metals is very high.

It is necessary to know the exact moment of the incorporation of the antioxidant since if the oxidation process is advanced, the antioxidant loses its capacity of action.

Among the main primary antioxidants include:

 1. Phenolic antioxidants

In this type, antioxidants are phenolic type hydrogen donors, and are able to effectively move an unpaired electron.

The main antioxidants of this type are:

  • Propyl gallate (E-310):  White crystalline powder used in food when other synthetic fat-soluble antioxidants are not suitable. It is not very soluble in water, and, in the presence of traces of iron, derived from food or from the equipment used in the processing, it gives rise to the appearance of unattractive, dark blue colors.
    Occasionally, Propyl Galato acts together with synthetic and natural antioxidants. It is important to bear in mind that it is a substance that is sensitive to high preparation temperatures.
  • Octyl gallate (E-311): Used as a synthetic antioxidant in fats and water, where it is sometimes included to prevent rancidity in oils.
  • Dodecyl gallate (E-312): Used as a synthetic antioxidant in fats and beverages, particularly to prevent rancidity in oils.

The most important technological property is the low resistance to heating. They are not very useful for protecting frying oils or sometimes foods that are subject to high cooking temperatures strong foods during their manufacturing, such as confectionery products or cookies. The low resistance to heat can be avoided by adding citric acid to the product. They are used, mixed with BHA (E 320) and BHT (E 321), for the protection of edible fats and oils.

Galactose, BHA and BHT were used together in oils, with the exception of olive oil. They are also used in canned and semi-preserved fish and processed cheese, pastry or confectionery, cookies.

2. Breakthrough phenols.

The main antioxidants of this type are:

  • Butyl-hydroxy-anisole (BHA, E-320): It is one of the most common antioxidants in human nutrition. Chemically, BHA is a mixture of two isomers: 2-tert-butyl-4-hydroxyanisole and 3-tert-butyl-4-hydroxyanisole. The second one is generally considered as a better antioxidant, and represents 90% of commercial BHA.

This antioxidant is effective primarily in animal fats and more discreetly in vegetable fats and oils. However, due to the chemical structure they present, they are extremely volatile at baking and frying temperatures.

  • Butyl-hydroxy-toluol (BHT, E-321): Together with BHA, they are the most used antioxidants in human nutrition. BHT (3,5-di-tert-butyl-4-hydroxytoluene) is an appropriate antioxidant for heat treatment, although it is not so stable.

It is commonly used in combination with BHA to provide increased antioxidant activity. It is also commonly used together with other antioxidants, such as propyl gallate and citric acid, for the stabilization of oils and high-fat foods.

Both BHA and BHT have a slight phenolic odor when used at high temperature for a prolonged period of time.

  • Terbutil hydroquinone (TBHQ, E-319): TBHQ is a white or beige powder that is frequently used in vegetable oils and animal fats. As an antioxidant, TBHQ is more effective in vegetable oils than BHA and BHT. It is heat-stable and very useful in the prevention of oxidation of frying oils.
    Like the BHA and the BHT, there are indications that in high doses it can be harmful to health, which is why the European Food Safety Authority (EFSA) has banned its use in Europe and the Food and Drug Administration of the United States (FDA) has established certain limits for its use in human nutrition.
  • Tocopherols (E-306): is the antioxidant extracted from nature most common in the food industry.
    It is completely fat-soluble and does not alter the organoleptic properties of the food. It is also safe, effective and easy to incorporate.
    The tocopherols are formed by four isomers (Alpha, Beta, Gamma and Delta) with different antioxidant and vitamin activities. After several exhaustive studies, it has been shown that the main antioxidant activity is produced by the Gamma and Delta isomers.

In products formed by structures with double bonds more input of tocopherols is needed, since the unsaturated substances are more sensitive to oxidation.

So far we have analyzed the primary antioxidants (type I), in another article, we will complete this analysis with the development of secondary antioxidants (type II).

Main nutrients in the diet for dogs and cats

Nutrition is the sum of processes by which an animal ingests and uses all the substances required for its maintenance, growth, production or reproduction. Through food, the living being satisfies its nutritional requirements, since it ingests the necessary nutrients for its conservation and growth. There are 6 different groups of nutrients:

Group of nutrients


An essential nutrient that can be considered as food. It is composed of suspended mineral particles called electrolytes. An animal that feeds on dry feed will take approximately 2.5 times the weight of the food it receives. However, in diets with high moisture content, the needs will decrease considerably.


A chemical substance that is part of the structure of cell membranes and is the essential constituent of living cells. It is formed by different chains of amino acids which  are classified as either essential or not essential. The essential amino acids are those that must be incorporated into the diets since the animal is not capable of synthesizing them itself. These are: arginine, histidine, isoleucine, leucine, lysine, methionine, fenialamine, threonine, tryptophan and valine.

Although taurine is not an amino acid itself, it is related to the functioning of the retina. Cats do not synthesize it in sufficient quantities, so they need an extra supply.


They are formed by Carbon, Oxygen and Hydrogen, forming covalent bonds that are difficult to break. Thanks to the oxidative phosphorylation of carbohydrates we obtain energy. They are present in two forms: glucose as an immediate energy supply and glycogen as a reserve. It is important to emphasize within this group, that fiber, although it is not a nutrient in itself, has a vital importance for the physiological functioning of the digestive system.

Fats or lipids

These nutrients constitute the main energy contribution in the food. Its main functions are:

  • to provide essential fatty acids
  • to transport the liposoluble vitamins
  • to give palatability and texture to food
  • to facilitate the swallowing of the food bolus.

Note  that the long-chain polyunsaturated fatty acids such as OMEGA-3 and OMEGA-6 are of major importance during pregnancy and lactation. They act on the immune, nervous, and cardiovascular systems and in general they improve the metabolism of the animal helping it acquire over time a shiny, firm and healthy coat.

The amount of fat contained in a diet must be directly proportional to the diet’s protein content. That is, the amount of fat can be increased if, in parallel, the protein content is increased.


They are organic substances that promote correct physiological functioning. According to their solubility capacity they are classified as water-soluble or liposoluble.

The water-soluble vitamins are only soluble in water, so they do not accumulate in the tissues, preventing hypervitaminosis states. The main ones are vitamin C and different B vitamins.

Imbalances in liposoluble vitamin produce important diseases. The lack of vitamin A or retinol is related to vision problems and decreased vitamin K intake with healing problems. A deficit of vitamin D produces alterations at bone level and the lack of vitamin E or tocopherols causes muscular dystrophy and erythrocyte hemolysis.


They are organic substances necessary for the growth and physiological maintenance of the animal. Their functions include structural capacity in bone and cartilage tissue, the regulation of neuromuscular transmission and catalytic activity as members of enzymes.

The most important minerals in animal feed is Calcium which helps maintain healthy bones and teeth, Sodium and Chlorine which regulate body fluids and Phosphorus which contributes to cellular and muscular function.

Nutritional Evaluation

Once the characteristics of each nutrient have been analyzed, the nutritional evaluation that will provide the necessary amount of nutrients for dogs and cats can be done. For this, we have to take into account several factors such as: height, race, sedentary lifestyle, maternity, lactation …

The nutritional evaluation is a process that is carried out in two parts:

  1. Evaluation of selection in each animal. Based on this selection, pets that are healthy and have no risk factors do not need an additional nutritional assessment.
  2. Extended evaluation. It is performed when one or more risk factors related to nutrition are found, or when the presence of these factors is suspected based on the selection evaluation.

There are several methods to determine the nutritional needs in an animal’s diet.

First, the final product is analyzed in the laboratory, this way the percentages of humidity, ash, proteins, fats, fiber and nitrogen-free extract will be determined, and from these data we will be able to obtain the maximum and minimum levels of each nutrient.

The second method will be done following the recommendations of the different agencies responsible for the study of animal nutrition. These standards will determine if a food is “complete and balanced”.

The minimum levels required vary, depending on the period that the animal is in; in case of growth or reproduction, the needs will be greater than in a maintenance period.

One way to know the nutrients in food is through the “Nutritional Guidelines for Complete and Complementary Food for Dogs and Cats” which will help producers of dog and cat feed to produce nutritionally balanced foods, in addition to meeting the requirements of the EU legislation on animal nutrition, both for adult animals, growing animals and in the breeding period, in addition to assessing the nutritional capacity of these foods.

These guidelines should be as rigorous as possible and will be based on up-to-date scientific studies. They also facilitate the evaluation of food, and cooperation between manufacturers, professionals responsible for animal care and competent authorities.

Therefore, we should always be sure that the food we are giving an animal is rich in minimal nutrients. The recommended daily allowance should satisfy all nutritional needs for both dogs and cats.

Importance of food safety

We can define food safety as the set of conditions and measures necessary during the production, storage, distribution, and preparation of food to ensure that its consumption does not represent a risk to the health of people due to physical, microbiological (bacteria , viruses, parasites) or harmful chemical substances, which can cause diseases of an infectious or toxic nature, whose manifestations range from gastrointestinal problems to long-term chronic diseases such as cancer.
Thus, food safety covers all the links of the food chain, from animal feed and production to sale to the consumer, through to processing, storage, transport, import and export.

Food safety when preparing food

Pathogens transmitted through food are imperceptible. At first sight, a food may seem innocuous and can still contain pathogens, be it bacteria, viruses or parasites that cause diseases.

The first step before cooking is to wash your hands with warm water and soap and clean the surfaces that are going to be in contact with the food. In addition, it is recommended to use different knives and cutting boards for each type of product. Contamination spreads when bacteria spread from one food product to another. This is especially common when preparing raw meats, fish, seafood and eggs.

It is important to take into account the optimum cooking temperatures, for this purpose a thermometer placed at different stages of the food is used, for example, to avoid consuming eggs cooked below 71.1 ° C. An example is Spanish omelette which,  due to its preparation, can reach different temperatures in different stages.

Finally, the product must be refrigerated as soon as possible, especially if the temperature is above 25 ° C, in which case the food  should not be out for more than an hour. Cold temperatures delay the growth of dangerous bacteria. Keeping the refrigerator at a constant temperature of 4.4 ° C or lower is one of the most effective ways to reduce the risk of diseases caused by bacteria in food.
You should never thaw food at room temperature. It is recommended to do it inside the refrigerator. If it is done in cold water or in the microwave oven, it should be cooked immediately.

Legislation on food safety

To ensure food safety, the European Union has adopted legislation that contains a series of rules that apply to the production, processing and introduction of food products in markets, with food safety being the primary responsibility of food companies and animal food and feed companies, while the competent authorities are responsible for monitoring, verifying and demanding compliance with this requirement through national systems of supervision and control in all the stages mentioned above. Thus, the member states of the European Union are also obliged to establish rules regarding the measures and sanctions applicable in case of violation of the legislation on food and feed, which must be effective, proportionate and dissuasive.

Particularly relevant in terms of food safety is the European Food Safety Authority (EFSA), which is the independent scientific body of the EU that assesses the risks present in food, a mission that it carries out through the compilation of all scientific evidence at a given moment about a specific risk in a food.

With all that information, EFSA deduces a toxicological reference value, also called guide value, based on health. For substances with a chronic effect, such as pollutants, this value is the maximum amount of substance to which the population can be exposed through the consumption of food, and from all other possible sources, throughout life in a safe way , that is, without presenting the toxicological effects of the substance. In the case of microbiological risks, EFSA evaluates the risk of exposure to certain pathogens for humans, as well as their toxins.

In its risk assessments, and based on data from the different substances or microorganisms present in the food and the consumption data of those foods of the diet at the European level, EFSA identifies the foods that pose the greatest risk for the exposure to said risk. for the general population, as well as for certain specific groups of the population, called vulnerable groups (children, pregnant women, the elderly, vegetarians, etc.).

On this scientific basis, experts from the different Working Groups of the Commission propose and discuss measures to avoid or reduce the exposure of European consumers to different food risks. This is what is known as food risk management, which is nothing more than evaluating the different options to protect consumers from food risks and which results in the establishment of the relevant legislation to achieve it.

While it is true that scientific data are very important when preparing legislation, other legitimate interests must also be taken into account when making decisions, such as economic, social or cultural interests. All this is contemplated in the process of elaboration of the legislation with the objective of avoiding that the legislation of alimentary security can get to constitute a true impediment to the international trade of the foods, and can endanger this economic sector or cause the shortage of a food in a country or region.

When discussing the risk management measures to be applied, it is necessary to weigh the benefit for the consumers (protection of the whole population or of specific groups) against the economic damage that can be caused in the commercialization of food.

Therefore we can conclude that the European legislation on food safety and food safety and in particular the regulations on food additives, is constantly evolving due to new scientific data, and thus, it is important to constantly keep updated in the novelties in legislative matter to be able to adapt to the new norms in alimentary production.

Physical properties of oils and fats

The analysis of the physical properties of oils and fats allows us to understand the behavior and characteristics of these elements, as well as their differences. For this, will be analyzed:

  • The crystallization
  • The melting point
  • The viscosity
  • The refractive index
  • The density
  • The solubility
  • The plasticity
  • The emulsifying capacity

Here we provide more detail on each of these.


Fats differ from oils in their degree of solidification at room temperature, since in these conditions the oils are in a liquid state (not crystallized) while the fats are in the solid (crystallized) state.

The proportion of crystals in fats have great importance in determining the physical properties of a product. Fats are considered solid when they have at least 10% of their crystallized components.

The fat crystals have a size between 0.1 and 0.5 μm and can occasionally reach up to 100 μm. The crystals are maintained by Van der Waalls forces forming a three-dimensional network that provides rigidity to the product.

An important feature of fat is its crystalline polymorphism, since mono-di and triglyceride crystallize in different crystalline forms (α, β, β’).

Form α (vitreous state)

  • Appears when the fat solidifies by a quick method.
  • The crystals formed are of the hexagonal type and are organized randomly in space.

Form β

  • It occurs when the cooling is slow or if the tempering is carried out at a temperature slightly below the melting point, this form being the most stable of all.
  • In the β form, tricyclic crystals are formed oriented in the same direction.
  • The β form is typical of palm oil, peanut, corn, coconut, sunflower, olive and lard.

Form β’

  • It is produced from the tempering above the melting point of the α form.
  • In the β-form, orthorhombic crystals are formed which are oriented in opposite directions.
  • The β’form is typical of modified partial cottonseed oil, fats, fats and modified lard.

Both α, β and β’form have a melting point, an X-ray diffusion pattern and a refractive index.

The more double bond there is, the crystallization with which it tends to be liquid is hindered.

Melting point

The melting point of a fat corresponds to the melting point of the β form, which is the most stable polymorphic form and is the temperature at which all the solids melt.

When short chain or unsaturated acids are present, the melting point is reduced.

The melting point is of great importance in the processing of animal fats.

The melting points of pure fats are very precise, but since fats or oils are made up of a mixture of lipids with different melting points we have to refer to the melting zone, which is defined as the melting point of the component of fat that melts at a higher temperature.


The viscosity of a fat is due to the internal friction between the lipids that constitute it. It is generally high due to the high number of molecules that make up a fat.

By increasing the degree of unsaturation the viscosity decreases, and when the length of the chain increases the fatty acids components also increases the viscosity.

Refractive index

The refractive index of a substance is defined as the ratio between the speed of light in air and in matter (oil or fat) that is analyzed.

Increasing the degree of unsaturation increases the refractive index, and when the length of the chain increases the refractive index also increases, that is why it is used to control the hydrogenation process.

As the temperature increases, the refractive index decreases.

The refractive index is characteristic of each oil and fat, which helps us to perform a quality control on them.


This physical property is of great importance when it comes to designing equipment to process grease.

Density decreases when fats dilate when going from solid to liquid

When the fats melt, their volume increases and therefore the density decreases.

For the control of percentages of solid and liquid in commercial fat, dilatometric curves are used.


Solubility has great relevance in the processing of fats.

Fats are fully soluble apolar solvents (benzene, hexane …).

Except for phospholipids, they are completely insoluble in polar solvents (water, acetonitrile). They are partially soluble in solvents of intermediate polarity (alcohol, acetone)

The solubility of fats in organic solvents decreases with increasing chain length and degree of saturation.

Phospholipids can interact with water because the phosphoric acid and the alcohols that compose them have hydrophilic groups.

Generally the surface tension increases with the length of the chain and decreases with temperature. Surface tension and interfacial tension decrease with ease with the use of surfactant agents such as monoglycerides and phospholipids.


It is the property that has a body to preserve its shape by resisting a certain pressure.

The plasticity of a fat is caused by the presence of a three-dimensional network of crystals inside which liquid fat is immobilized.

For a grease to be plastic and extensible there must be a ratio between the solid and liquid part (20 -40% solid state fat), the nets must not be tight and their crystals must be in α form.

The plastic fats act as a solid until the deforming forces that are applied break the crystal lattice and the grease passes to behave like a viscous liquid and therefore can be smeared.

Emulsifying capacity

The emulsifying capacity is the capacity in the water/oil interface allowing the formation of emulsion.

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Regulation of antioxidants in Animal Nutrition

One of the main barriers facing producers today is to grant antioxidants the name of feed additives or veterinary drugs. This classification is important since, if veterinary medicines are considered, more exhaustive controls should be passed than in the case of additives.

According to Regulation (EC) No 1831/2003, antioxidants in animal nutrition will be considered “technological additives” defining antioxidants as: “substances that prolong the shelf life of feed and feed materials, protecting them against spoilage caused by oxidation. ”
The same regulation establishes the comunity procedure for the authorization of the commercialization and use of additives for animal feed introduces the rules of surveillance and labeling of additives and premixtures for animal feed. It therefore establishes the basis to guarantee a high level of protection of human health, animal health and welfare, and the environment, as well as the interests of users and consumers, on additives intended for animal feed.

The Regulation establishes the difference between “food additive” which is basically considered as the substance that helps to promote good health of the animal and therefore have a good performance, compared to “veterinary products” that serve more as a treatment of certain specific disorders.

It is worth highlighting the importance that currently exists for many antioxidants, such as vitamins, but these are not based on any law that regulates or investigates the use of antioxidants, and the problem that could lead to excessive use is known of these antioxidants, leading to a prooxidation, with an opposite effect to the desired one.

An additive for animal feed is considered to be any substance, micro-organism and preparation other than feed materials and premixtures, which are added intentionally to feed or water, in order to carry out in particular one or more of the following functions:

  • positively influence the characteristics of the feed;
  • positively influence the characteristics of animal products;
  • favorably influence the color of ornamental birds and fish;
  • satisfy the nutritional needs of the animals;
  • positively influence the environmental impact of animal production;
  • positively influence the production, activity or welfare of the animals, especially acting on the  gastrointestinal flora or the digestibility of the feed, or have a coccidiostatic or histomonostatic effect.

As stated in Regulation (EC) 1831/2003 of the European Union on the use of additives in animal feed, livestock production has an important place in the agriculture of the Community, and part of the benefits of this livestock production are due to the use of safe and good quality feed, a good feeding of the animals, which will ultimately affect the health of the citizens.

Therefore, to increase the protection of human health, animal health and the environment, the additives that will be used in animal feed must follow a series of safety assessments according to what the European Union procedure indicates, prior to marketing, use or transformation.

In this way, only authorized additives as established in this regulation may be used or transformed for use in animal feed. In addition, the categories of the additives must be defined in order to be able to evaluate their authorization of use more adequately.

The rules that refer to the request for this authorization of additives for animal feed must take into account a series of requirements:

  • Food-producing animals
  • Other types of animals

Together with this standard, the evaluation of the European Food Safety Authority (Regulation n° 178/2002) will have to be considered, since in these evaluations the waste presented by the food will be valued (Maximum Residue Limit, or MRL).

But this risk assessment cannot offer all the information necessary for risk management and the subsequent authorization to use an additive. In addition, other factors of a sociological, economic and environmental nature must be taken into account, as well as the viability of the controls and the benefit to the animals.

The Commission responsible for the authorization of feed additives and their conditions of use must inform about their maintenance and must publish a register of authorized additives. In addition, the holder of said authorization will have to follow a surveillance plan in accordance with the traceability requirements dictated by food legislation, after the product is marketed, to assess any effect that mixing these additives in the feed may cause on human health, animal health or environment.

The authorization will be for a limited time and in this way a periodic review of the additives will be favored.

A record of authorized feed additives should be established, including specific information on the products, in addition to their detection method.

A detailed labeling of the products allows the end user to choose with full knowledge of the cause and also reduces the obstacles to its commercialization, as well as facilitating the fairness of the transactions.

How to request the authorization of an additive

The application must be submitted to the Commission in charge of its management, for the use of additives for animal feed. This Commission shall inform the Member States thereof and transfer it to the European Food Safety Authority.

This request must be presented with the following information and documentation:

  1. The name and address of the applicant
  2. The identification of the feed additive, a proposal to classify it by category and group, and its specific data, including, the degree of purity when applicable.
  3. A description of the method of production and manufacturing and the expected uses of the additive for animal feed, the method of analysis of the feed additive according to its intended use, and, where appropriate, the method of analysis used to determine the level of the residues of the feed additive, or its metabolites, in food
  4. A copy of the studies carried out and any other material available to demonstrate that the feed additive meets the established criteria
  5. The proposed conditions for the marketing of the feed additive, including labeling requirements and, where applicable, specific conditions of use and handling (including known incompatibilities), levels of use in supplementary feed and animal species and categories to which the feed additive is intended
  6. A written declaration stating that the applicant has sent three samples directly to the aforementioned Community reference laboratory according to a set of established requirements.
  7. A proposal for follow-up to the commercialization
  8. A summary of the file that includes the information provided
  9. Information on any authorization granted under the applicable legislation, for additives falling within the scope of Community legislation on the marketing of products that consist of, contain or are produced from GMOs.

After submitting the application, the Authority will issue an opinion in a period of approximately six months, in which they will have reviewed and verified the information and documentation presented, and will verify the report in the reference laboratory.

Methods to determine oxidative stability

The autooxidation 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 autooxidation 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: initiation, propagation and 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 Methods

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.

They 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.
  • p-Anisidine: Quantifies oxidation by-products such as high molecular weight carbonyl compounds. An oxidized and deodorized oil would be detected by this analysis.
  • TBA: 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.
  • Acidity: 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

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: it 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: it 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.

Packaging and food preservation

Food is an ideal medium for the growth of microorganisms. Therefore, by inhibiting the development of these, we can increase the shelf life of food.

There are many causes that negatively influence the quality of food, either by intrinsic factors of the food, such as its nutrient content, water availability, pH, etc., or by extrinsic factors such as temperature of storage, relative humidity, exposure to sunlight and air, handling and processing of raw materials, etc.

The main objective of food preservation is to maintain a product in perfect hygienic conditions and to protect its rheological and organoleptic qualities (Casp and Abril, 2003).
The processes of food preservation allow us to obtain safe products of high quality at a reasonable price. The permanent increase in the demands of consumers in terms of quality and prolongation of the useful life of the food, causes continuous changes in the way in which the food is produced, distributed, stored and sold. The food industry is constantly searching for new methods that are less aggressive with food, with lower energy consumption, and more effective against pathogenic microorganisms.

Among the different procedures that guarantee these expectations, we can highlight its relevance to packaging.


Packaging is a conservation method that protects food from light, moisture and other environmental contaminants. For a correct packaging process, the following factors must be taken into account:

  •  Storage: capacity to be stacked and transported, control of quantity produced, conservation of small products,
  • Protection: against deterioration, leakage, breakage, dehydration, contamination, theft and alteration.    Physical protection against shock, vibration, compression, temperature, etc.  Barrier protection against     oxygen, water vapor, dust, bacteria, etc.
  •  Information: identification of the product, description of use or preparation, warning about risks derived from  improper use, list of ingredients, nutritional data and price, etc.
  • Promotion: marketing tool to differentiate the product from similar ones and attract attention in shops and supermarkets using, for example, brands, colors, illustrations and forms.
  • Transport: greater ease and safety to move products from the manufacturer to the warehouse and the vendors (tertiary containers) and even the consumer (primary packaging).

Currently we can find a variety of materials for the manufacture of containers, with different gas permeabilities, with variable resistances and permissiveness to light (transparent, translucent, opaque), but also, there are elements that allow us to know if temperature has been constant during storage or if there have been breakages of the cold chain, as well as the concentration and composition of the gas inside (Rodríguez, 2004).

Due to its versatility in shape and size, being light and hygienic, the most used materials are synthetic plastics. However, because they are products that do not decompose, they are a cause of environmental pollution. Therefore, in recent years new types of less polluting or easy to recycle materials have been developed, which are called biodegradable packaging.

The biodegradable containers come from renewable sources and  many of them are characterized  as being edible. They are applied as a barrier for microorganisms and to improve sensory properties such as appearance, color, brightness and transparency.

Currently the container, in addition to fulfilling its basic functions of containment, protection and information, is becoming a medium that performs sophisticated interactions with its content and in a registry of important information for both the consumer and the intermediaries of the chain of value, thus creating the concept of active packaging.

It is considered to be an active element when there is another function other than providing an inert barrier against external conditions. They belong to this group of products when food components or some material  are used as an indicator of the history and quality of the product. There are two mechanisms of action to create this type of packaging: introduce the active element inside the container together with the food or that it is part of the food itself.

A way to achieve active packaging by incorporating natural antioxidants such as tocopherols, which allows to increase the shelf life of foods by inhibiting or delaying the oxidation of lipids or other compounds (Quezada-Gallo, 2009).

Among the active packaging, the concept of intelligent packaging stands out. It is a packaging system capable of collecting and processing information from the environment in order to transmit it to the consumer (Aguirre, et al, S / F) Its objective is to ensure the quality of the product by monitoring the processes that alter the food

Another method of food preservation that has stood out in recent years, is packaging in modified atmosphere. It consists of the alteration of the environmental gases to reduce the microbial growth and the speed of the internal chemical reactions.

In commercial practice, oxygen reduction and increase of carbon dioxide and / or nitrogen are usually used. Carbon dioxide acts as a destructive agent of bacteria and fungi and reduces the multiplication of pathogenic microorganisms.

In the process of packaging in modified atmospheres it is not necessary, in general, to maintain the composition of the gas throughout the storage, so it is more practical and economical.

When calculating the amount of modified gases, it must be kept in mind that the effect is not the same for all products and working conditions. It varies according to the composition, characteristics and sanitary status of the food to be conserved, the composition of the atmosphere and the storage temperature, as well as the packaging materials and packaging technology. (R. Catalá and R. Gavara, 2001)

Nowadays, packaging is vital in the marketing of food, since in addition to providing better conservation, longer life of the food and information for the consumer, they must produce a visual impact that makes them differentiate themselves from similar products to be chosen by the final consumer (Cruz, 2006).

A suitable container should avoid contamination of the food by preventing the passage of outside substances. In some cases, the packaging can cause alterations to the taste, smell or texture of food and be harmful to health, so it is necessary to control the materials with which it has been manufactured


The main advantages that good packaging brings to the quality and conservation of food are:

  • To preserve the organoleptic properties of the food and, therefore, its quality.
  • Lengthen the shelf life of the product.
  • Slow down enzymatic and microbial reactions.
  • Decrease weight loss due to evaporation.
  • Enable a more hygienic transport and storage.
  • Eliminate dripping and unpleasant odors.
  • Improve the final product presentation

Effects of Antioxidants in Poultry

A large number of articles highlight the importance of antioxidants in animal feed. Its use is justified in order to avoid the appearance of “oxidative stress”, which can cause losses in the productive performance, as well as losses in both nutritional and organoleptic quality of the products derived from them.

What is oxidative stress?

Oxidative stress is the imbalance between the endogenous generation of free radicals and the antioxidant defense system of the organism (Halliwell and Gutteridge, 1999).

There are a number of endogenous antioxidants,  produced by the body itself, to fight against these free radicals, but at a certain time it is necessary to add exogenous antioxidants in order to maintain the physiological balance between an excessive production of free radicals and an adequate amount of antioxidants in the body.

Therefore, to diminish these negative effects on the organism and the meat, several studies have confirmed the efficacy of antioxidants both “in vivo” and post-mortem.

In this article we will evaluate the consequences of the use of antioxidants, both during the animal’s life and after its slaughter, as well as in the products obtained.

It is important to understand when an extra addition of exogenous antioxidants is necessary in the diet, making it  crucial to identify the factors that cause an imbalance in the organism of birds, among which we can highlight:

  • Incorrect temperatures on the farm, in addition to high levels of ammonia and carbon dioxide due to poor ventilation.
  • Situations that cause stress to the animal
  • Toxics or heavy metals, some medications, even excesses of Vitamin A
  • Feeding with oxidized fats.
  • Mycotoxins

Effects of Antioxidants in Vivo:

During the life of the animal, there are several factors that could affect its final production, such as pathological, nutritional, physiological or environmental causes that cause oxidative stress and reduce the performance of animals.

It is considered that the components of the feeding will have a modulating effect in the maintenance of this balance. As indicated by a large number of studies (Fellenberg and Speisky, 2006, Salami et al., 2015), which highlight the importance of the use of antioxidants in broilers and poultry, as well as in breeding animals.

Antioxidants and Fertility:

Sperm have a high amount of polyunsaturated fatty acids, which will be very susceptible to oxidative stress, so an antioxidant protection will be essential for motility and fertility.

Therefore, diets rich in antioxidants would help increase the level of antioxidants and thus improve motility and fertility.

Embryonic development:

The tissues of the embryo have a large amount of polyunsaturated fatty acids, hence their ability to oxidize and their need for antioxidant protection. Many of them are released by the mother, during egg laying, and thanks to the food provided, the amount of antioxidants such as Vitamin E or Selenium may be higher. Others will be synthesized by the embryo’s own tissues.

The amount of lipids and the concentration of antioxidants will determine the vulnerability of the embryo to free radicals.

Diets with supplemented by  antioxidants would help to increase the concentrations in the chicken and decrease its susceptibility to lipid peroxidation

Effects of Postmortem Antioxidants:

In addition to these “in vivo” effects, antioxidants can also have postmortem effects, in the meat obtained from the animal as well as in the egg. These points are developed below:

Effects on Meat

Among the improvements in the quality of the meat due to an increase in in vivo antioxidant supplementation we could highlight:
– Improvements in color and oxidative stability of lipids.
– Reduction of odor and rancid taste, both raw and cooked.
– Increase of the CRA (Water retention capacity) “prevention of fluid losses”

Effects on the Egg

At present, the diet provided to birds is rich in n-3 fatty acids, since they are considered to be of great importance for growth, development, as well as being beneficial for human health.

By increasing these fatty acids in the diet, the amount of fatty acids in the egg also increases, which will cause it to deteriorate faster. In order to reduce this deterioration, it was observed that antioxidants, especially Vitamin E, prevented the degradation of fatty acid even after several days of storage.

We can conclude that lipid oxidation in poultry production will later affect the quality and deterioration of products destined for human consumption. In order to reduce these losses in the product, a series of antioxidants suitable for each stage must be used, taking into account the different factors that catalyze the start of the same.