FIELD OF THE INVENTION

The present invention refers to the use of a composition for distributing antioxidants to an animal feed or feed raw material, wherein said composition comprises: a) a first antioxidant

b) a second antioxidant

c) a glycerol ester composition comprising glycerol esters of propionic and/or butyric acid, and wherein said first antioxidant and said second antioxidant are not identical compounds.

BACKGROUND OF THE INVENTION

Animal production is challenged more than ever with changing legislation, circumstances and conditions. A safe and healthy production of these products is extremely important. From an economical point of view it is also important to remain or even improve the performance in animal production . The quality of ingredients and final products is a key, preventing spoilage due to oxidation or microbial contamination.

The oxidation process in fats and oils is usually described as“autoxidation”. It is a chemical process which is fairly complex with many chemical reactions. This destructive process is initiated and catalyzed by various parameters such as metal ions, increased temperature, moisture, light and oxygen as well as by lipoxygenases from vegetable or microbial origin. The classical route of autoxidation depends on the production of free radicals formed from fatty acids by hydrolytic oxidation. The resulting radical combines with oxygen to form a free radical. This in turn is converted into a hydroperoxide and another free radical and the reaction propagates automatically.

The open porous structure of many products and the high temperatures during process increase the risk for autoxidation. The presence of catalysts, such as copper or iron, also leads to a more rapid oxidation of fats. Heat processing also activates other oxidative catalysts, such as haemoproteins containing iron. Furthermore, fats or oils are often sprayed on the outside of pellets and the large surface area brings the fat in good contact with oxygen. The first products of oxidation of a fat are odorless, tasteless radicals, peroxides and hydroperoxides. Radicals react very rapidly with oxygen forming peroxides and

hydroperoxides, which further breakdown and produce hydrocarbons, aldehydes, ketones, alcohols and finally short chain fatty acids. The production of these end products is called rancidity. The undesirable odors in rancid products can be caused by very small quantities of only a few ppm of aldehydes or ketones. This influences the palatability of the feed in a negative way, which in turn can have severe effects for animal growth and performance.

Further on as oxidation occurs, it generates heat, which represents a loss of metabolizable energy available to the animal. The metabolizable energy increases as the degree of unsaturation in the fat increases. The susceptibility of the fat to oxidation also increases with the degree of unsaturation. Therefore as the fat oxidizes not only is the energy being lost but also the most available energy, represented by the unsaturated fats, is lost first. The by products of the autoxidation process include the production of low molecular weight components. They have no nutritional value and can even be toxic to the animal. Peroxides and other intermediate components of the autoxidation can interact with and destroy fat- soluble vitamins and even reduce protein digestibility.

Some studies have also shown that feeding of oxidized fats to broilers influenced functional changes in the gastrointestinal system (JJ Dibner et al, 1996). There were also transient effects on the intestinal microflora and efficacy of the gut associated immune system was impaired.

The increasing requirement for improved food and feedstuffs has encouraged the

manufacturers to introduce more specific ingredients and advanced processing techniques. Animal feeds have to fulfil the requirements of the nutritional, health, and welfare needs for each kind of domestic animal to optimize growth, reproduction, milk production, egg production etc. The tendency in most types of feed is an increase in energy via fat addition in the mixer and/or via a coating on the outside of the pellets after the dryer in case of high energy level feeds. The added fat in poultry feeds can go up to 10%, whereas in premium petfood diets and feed for aquaculture lipid contents of respectively 25% and 40% are reached. There is also a trend towards increased temperatures involved in the manufacturing processes, such as in extrusion, expanding and pelleting, to obtain highly digestible diets with reduced microbial contamination. Animal production is no longer seen as just a source for proteins and fats. Now a days it makes a part of the total food chain. The consumer more and more defines the characteristics of the final product he wants to eat. Feed is turned into food. The quality of raw material and final feeds is essential. It is important in human nutrition to minimize the intake of oxidized materials as this may stimulate peroxidation in living tissues. This has important consequences for the onset of many diseases.

Including and proper use of specific antioxidant systems is a vital part of good feed production practices. The prevention of autoxidation is not simply adding an antioxidant to the feed or food. Avoiding oxidation of ingredients in an early phase is a critical parameter. It is therefore essential that fats, oils and lipid-rich raw material products are treated with antioxidants already before storage or latest before production of the feed. The more intermediate components of the autoxidation are formed the more other nutrients such as proteins, amino acids, carotenoids, metabolizable energy and vitamins can be affected during the manufacture and storage of the high temperature processed feeds. Additional antioxidant is also included in the diet or via the premix in order to preserve the natural oils, fats and flavors present within cereals and raw materials and to obtain a well-stabilized feed.

Antioxidants for food and feed are classified as either primary or secondary antioxidants. Primary antioxidants terminate the free-radical chain reaction by donating hydrogen or electrons to free radicals and converting them into more stable products. They may also function by addition in reactions with the lipid radicals, forming lipid-antioxidant complexes. Both hindered phenolic (e.g. BHA, BHT, TBHQ and tocopherols) and polyhydroxyphenolic (e.g. gallates) antioxidants belong to this group.

Secondary or synergistic antioxidants can be broadly classified as oxygen scavengers and chelators. Synergists function by various mechanisms. They may act as hydrogen donors to the phenoxy radical, thereby regenerating the primary antioxidant. Hence phenolic antioxidants can be used at lower levels if a synergist is added simultaneously to the product. Synergists also provide an acidic medium that improves the stability of primary antioxidants.

Oxygen scavengers such as ascorbic acid, ascorbyl palmitate and sulfites react with free oxygen and remove it in a closed system. They may also act as synergists with primary antioxidants. Chelators like ethylenediaminetetraacetic acid, citric acid and phosphates are not antioxidants, but they are highly effective as synergists with both primary antioxidants and oxygen scavengers. An unshared pair of electrons in their molecular structure promotes the chelating action. They form stable complexes with prooxidant metals such as iron and copper and raise the energy of activation of the initiation reactions considerably.

Combining antioxidants in order to achieve synergistic effects is quite complex. A specific combination of antioxidants can give a synergetic effect when used together under certain conditions. It could be for example two primary antioxidants, a primary and a secondary molecule or more than one primary antioxidant and one or more secondary antioxidants working together. Which combination works best depends on which type of fat, oil, protein meal etc. should be protected and at which concentration ratios the antioxidant molecules are added to the system.

As already happened in human foods and since a few years in petfoods, antioxidant molecules which are not approved for food application will no longer be tolerated. A similar approach is going on for example in aquaculture, where fish meal and fish oil are the major ingredients of feed. These ingredients have traditionally been stabilized with ethoxyquin, which has now been banned in the EU and is being phased out. Moreover, the maximal permitted level of single antioxidants is often a limitation to how long the ensured shelf life of a feed product can be. It has become obvious that single antioxidants no longer fulfil today’s requirements.

Therefore antioxidant systems based on synergistic combinations of antioxidants, natural antioxidants and chelators are being more and more used to avoid spoilage of feed due to autoxidation. Further, there are indications that different antioxidant moleculs will influence the generation of different oxidative components, as shown by C. Liang et al, 1998, using an oxidizing model system. For example, combinations based on BHA ((1,1 -dimethylethyl)-4- methoxyphenol) and BHT (2,6-bis( 1 , 1 -dimethylethyl)-4-methy lphenol) are more effective in the prevention of oxidation of animal protein meals and animal fats whereas combinations with BHA and gallates give a better protection of vegetable oils and marine oils, products with a higher level of unsaturated fatty acids.

So, it is desirable to be able to design different combinations of antioxidants for different applications, in order to achieve the best synergistic antioxidative effect possible. However, certain combinations of antioxidants are difficult to dissolve together in the same system. The reason for this can for example be that one of the antioxidants is only soluble in non-polar solvents, such as rapeseed oil, and the other antioxidant is only soluble in more polar solvent systems, such as monoethyleneglycol or polyethyleneglycol/water. Combinations like BHT/propyl gallate is neither dissolved in rapeseed oil nor in polyethyleneglycol/water and has until now not been possible to use together in the same antioxidant system. It has now surprisingly been found that an antioxidant composition with for example BHT and propyl gallate can be dissolved in a glycerol ester composition comprising glycerol esters of propionic acid or butyric acid respectively, which are all fully approved as feed additives in the EU.

The use of polyethylene glycol/water for dissolving antioxidants for animal feed applications is no longer approved in the EU, which calls for alternatives where this solvent system has been used until now. The antioxidant combination BH A/propyl gallate/citric acid was previously dissolved using polyethylene glycol/water as a solvent system to obtain a low viscous liquid antioxidant product optimal in application. It has now been found that this antioxidant combination can be readily dissolved using a glycerol ester composition comprising glycerol esters of butyric acid.

DETAILED DESCRIPTION OF THE INVENTION

The feed manufacturing business is being challenged with new requirements on antioxidants used in animal feed and feed raw material, both from authorities and from consumers. The EU ban of ethoxyquin, a tendency at some petfood and aquatic feed producers to avoid BHA and the EU non-approval of polyethyleneglycol/water as solvent system for antioxidants in feed products are all examples of this. There is clearly a need for new, more specific and more synergistic combinations of antioxidants to be able to meet these requirements and to provide the animal production industry with antioxidant systems to avoid autoxidation and produce safe feeds.

The present invention refers to the use of a composition for distributing antioxidants to an animal feed or feed raw material, wherein said composition comprises: a) a first antioxidant

b) a second antioxidant

c) a glycerol ester composition comprising glycerol esters of propionic and/or butyric acid, and wherein said first antioxidant and said second antioxidant are not identical compounds.

The risk for oxidation of a feed or feed raw material increases as the fat content of the product increases and therefore it is of course important to protect fat-rich feeds or feed ingredients. Other nutrients, like proteins, carotenoids and fat soluble vitamins can get negatively affected by the intermediates formed in the process of autoxidation, so these ingredients are also important to protect from oxidation reactions.

According to one embodiment of the present invention the feed or feed raw material comprises a pet food, a pet food raw material, a protein meal, a protein meal raw material, a fat, an oil, fat soluble vitamins, carotenoids and/or a vitamin- and/or mineral premix.

As discussed above there are different kinds of antioxidants that have different modes of action, working alone or together with one or more other antioxidants.

It has become increasingly important to be able to design specific combinations of antioxidants for specific applications; this in order to achieve the most effective combination of two or more antioxidants working synergistically together. However, some combinations of antioxidants may be difficult or even impossible to dissolve together in any of the solvent systems currently used together with antioxidants. In some cases, one of the antioxidants in the system is only soluble in polar solvents, such as monoethyleneglycol or polyethyleneglycol, and another, second antioxidant, that needs to be dissolved together with the first antioxidant, is only soluble in non-polar solvents, such as rapeseed oil. For example, the combination of BHT and propyl gallate cannot be dissolved either in polar solvents or in non-polar solvents.

The present invention offers a solution to this problem by using a glycerol ester composition for dissolving two or more different antioxidants in the same system and distributing the antioxidant-glycerol ester composition to an animal feed or feed raw material. For example, the specific combination BHT/propyl gallate is readily dissolved according to the present invention, by using a glycerol ester composition comprising either glyceryl tributyrate or glyceryl tripropionate.

According to one embodiment of the present invention, the first antioxidant is only soluble in polar solvents and the second antioxidant is only soluble in non-polar solvents.

According to a preferred embodiment of the present invention, the composition used for distributing the antioxidants to the feed or feed raw material comprises: a) 0.1-35 parts per weight of said first antioxidant

b) 0.1-35 parts per weight of said second antioxidant

c) 10-90 parts per weight of said glycerol ester composition.

Glycerol esters of short chain fatty acids are very well suited to be used according to the present invention. Besides being fully approved as feed additives in the EU, many of these esters are today used to improve gut health in animals and to contribute to an improved animal performance. By using a glycerol ester composition according to the present invention, not only can antioxidants be distributed to a feed or feed raw material in a safe and convenient way, the glycerol esters can also, at the appropriate diet inclusion level, contribute to an improved gut health for the animal consuming the feed.

Glycerol ester compositions comprising butyric acid have proved to be particularly suitable for use according to the present invention.

According to a preferred embodiment of the present invention, a glycerol ester composition comprising 20-60 parts per weight of glyceryl monobutyrate, 5-40 parts per weight of glyceryl dibutyrate and 0.5-10 parts per weight of glyceryl tributyrate is used. In the context of the present invention, this glycerol ester composition is referred to as“mono-/dibutyrin”.

As discussed above, combining primary and secondary antioxidants in different ways is often necessary in order to achieve a satisfactory antioxidative effect. According to one embodiment of the present invention, the composition comprises at least one secondary or synergistic antioxidant, such as a chelating agent or an oxygen scavenger. According to a preferred embodiment of the present invention, said at least one secondary or synergistic antioxidant comprises citric acid.

The mono-/dibutyrin glycerol ester composition has proved to work very well for dissolving for example the antioxidant combination BHA/propyl gallate/citric acid. This combination of antioxidants has previously been dissolved using polyethylene/water, a solvent system that is no longer approved for use in feed by the EU.

According to a particularly preferred embodiment of the present invention, the (total) composition comprises 0.1-35 parts per weight of BHA, 0.1-35 parts per weight of propyl gallate, 0.1-25 parts per weight of citric acid, 5-50 parts per weight glyceryl monobutyrate, 1- 30 parts per weight of glyceryl dibutyrate and 0.1-10 parts per weight of glyceryl tributyrate.

Further on, according to another preferred embodiment of the present invention, a glycerol ester composition comprising 0.5-10 parts per weight of glyceryl monobutyrate, 1-30 parts per weight of glyceryl dibutyrate and 50-98.5 parts per weight of glyceryl tributyrate is used. In the context of the present invention, this glycerol ester composition is referred to as“tributyrin”. Using this glycerol ester composition according to the present invention, the antioxidant combination BHT/propyl gallate can be dissolved and distributed together to an animal feed. This combination of antioxidants has until now not been possible to dissolve together.

According to a particularly preferred embodiment of the present invention, the (total) composition comprises 0.1-35 parts per weight of BHT, 0.1-35 parts per weight of propyl gallate, 0.1-10 parts per weight glyceryl monobutyrate, 1-20 parts per weight of glyceryl dibutyrate and 30-90 parts per weight of glyceryl tributyrate.

According to still another embodiment of the present invention, a glycerol ester composition comprising 0.5-10 parts per weight of glyceryl monopropionin, 1-30 parts per weight of glyceryl dipropionin and 50-98.5 parts per weight of glyceryl tripropionin is used. In the context of the present invention, this glycerol ester composition is referred to as“tripropionin”.

The composition used according to the present invention is added to the animal feed or animal feed raw material at a concentration of 10-5000 ppm. In practice, regulations on maximum additions of antioxidants often set the limit for which concentration can be allowed in a particular feed. For example, for BHT and BHA this limit is 150 ppm alone or combined and for propyl gallate this limit is 100 ppm.

The antioxidant solution is often applied using a dosing system, spraying the solution on fresh raw material, into a fat/oil line in production, onto the feed during processing or on processed products.

The feed or feed raw material to which antioxidants have been distributed according to the present invention can be feed intended for consumption by companion animals, aquatic species like fish or prawn, poultry, swine, cattle, horses and/or sheep.

The susceptibility of a feed or feed raw material to future oxidation can be predicted using so- called oxidation stability tests. The oxidation stability of an oil, a fat or a feed could be ascertained reliably by periodically examining samples kept in actual environmental use conditions, using different oxidation level tests. However, this would be too time-consuming and would not be feasible because of practical reasons. Oxidation stability is usually assessed using accelerated tests, in which matrix oxidation is forced by exposure to heat, oxygen and air, among other factors. These accelerated tests are very useful to compare antioxidants and to assess raw material feeds with and without antioxidants. The results of these tests are expressed as induction times, i.e. the times needed for the oxidation process to move from a latent or slow phase onto a fast and accelerated phase.

One of the oxidation stability tests commonly used is the Oxipres™ method. The Oxipres™ method is based on oxygen consumption at high temperatures and pressures. In an Oxipres™ test, the sample is placed inside a hermetically closed vessel that is subjected to high oxygen pressures (> 5 bar) and temperatures of 90 to 120 °C. The system is connected to a transducer that monitors pressure changes inside the pressure vessel and sends signals which are recorded and plotted on a computer, where pressure is expressed over time.

Figure 1 shows an example of curves obtained with an Oxipres™ test. In this case, the sample is an animal fat protected with two different levels of an antioxidant and a control sample without antioxidant. As can be seen in the figure, the pressure rises early during the test, because of the increase in temperature, and subsequently stabilizes until it begins to decrease as a result of oxygen consumption during the oxidation process. The induction point for each sample is the inflection point where the pressure begins to drop at a given temperature.

The present invention is illustrated in the below Embodiment Examples, which are to be construed as merely illustrative and not limiting in any way:

— Example 1 illustrates the use of different compositions for dissolving BHA/propyl gallate/citric acid

— Example 2 illustrates the use of different compositions for dissolving BHT/propyl gallate.

— Example 3 illustrates the antioxidative efficacy of BHT/propyl gallate dissolved in tributyrin (according to the invention) on substrate poultry fat - comparison is made with existing product of BHT/BH A dissolved in rapeseed oil.

— Example 4 illustrates the antioxidative efficacy of BHT/propyl gallate dissolved in tributyrin (according to the invention) on substrate duck fat - comparison is made with existing product of BHT/BHA dissolved in rapeseed oil and with natural mixed tocopherols.

— Example 5 illustrates the antioxidative efficacy of BHT/propyl gallate dissolved in tributyrin (according to the invention) on substrate pore protein meal.

EMBODIMENT EXAMPLES

Example 1: Use of different compositions for dissolving BHA/propyl gallate/citric acid

A combination of the three antioxidants BHA (( 1 , 1-dimethy lethyl)-4-methoxyphenol), propyl gallate and citric acid was dissolved in three different solvent systems, according to Table 1 below, while thoroughly mixing.

Table 1.

The non-polar solvent rapeseed oil could not dissolve this specific combination of antioxidants. The more polar solvent system polyethylene glycol/water was able to dissolve the antioxidants. However, this solvent system is not approved by the EU anymore for use in animal feed.

Using a glycerol ester composition comprising glycerol esters of butyric acid, according to the present invention, resulted in a thorough solution that is EU-approved, non-viscous and easy to handle.

Example 2: Use of different compositions for dissolving BHT/propyl gallate

A combination of the antioxidants BHT(2,6-bis(l,l-dimethylethyl)-4-methylphenol) and propyl gallate was dissolved in three different solvent systems, according to Table 2 below, while thoroughly mixing.

Table 2.

In this case, neither the non-polar solvent rapeseed oil, nor the polar solvent system polyethylene glycol/water could dissolve the combination of BHT and propyl gallate. Both the experiments performed according to the present invention, using a glycerol ester composition comprising glycerol esters of butyric acid and glycerol esters of propionic acid respectively, resulted in a thorough solution that is EU-approved, non-viscous and easy to handle.

Example 3: Efficacy test of BHT/propvl gallate in tributyrin on substrate poultry fat comparison with existing product of BHT/BHA

An antioxidant efficacy test was performed with the antioxidant formulation 15% BHT and 5% propyl gallate, dissolved in tributyrin, according to the present invention, on fresh poultry fat. The antioxidant formulation was added at three different concentrations. The antioxidant combination BHT/propyl gallate is not soluble in solvents conventionally used for

antioxidants, instead a comparison was made with the combination 15% BHT and 5% BHA dissolved in rapeseed oil.

The oxidation of the poultry fat was measured using the Oxipres™ method described above. The temperature in this test was 100°C. As a result of the consumption of oxygen the pressure in the vessel will drop after some time. The induction point is defined as the time elapsed between placing the pressure vessel in the block heater and the inflection point or break point when the pressure starts to drop, at a given temperature (100°C in this case). The result of the experiments can be seen in Table 3 below.

Table 3.

comparison with existing products

The efficacy experiment described in Example 3 was repeated, but in this experiment the substrate was duck fat. The antioxidant combination BHT/propyl gallate dissolved in tributyrin (according to the invention) was tested and compared to the commonly used antioxidants BHT/BHA in rapeseed oil and natural tocopherols respectively. The result of the experiments can be seen in Table 4 below.

Table 4. Example 5: Efficacy test of BHT/propyl gallate in tributyrin on substrate pore protein meal The efficacy experiment described in Example 3 was repeated, but in this experiment the substrate was a pore protein meal. The antioxidant combination BHT/propyl gallate dissolved in tributyrin (according to the invention) was tested. The result of the experiments can be seen in Table 5 below.

Table 5.

The above embodiment examples shows the applicability of the present invention on different substrates and with different combinations of antioxidants. Using a composition according to the present invention has made it possible to use combinations of antioxidants that has until now not been possible to use together (BHT/propyl gallate). It has also been shown that using a composition according to the present invention has made it possible to use certain

combinations of antioxidants together that has until now only been possible to use together when dissolved in a solvent system that is no longer approved by the EU (BHA/propyl gallate/citric acid). The antioxidative efficacy when using a composition according to the present invention has been shown to be in line with compositions commonly used today. This together with the benefits mentioned above makes the use of a composition according to the present invention an excellent option for distributing antioxidants to an animal feed or feed raw material.