In recent years, we can observe an increase in the demand for proteins in animal feed application, due to the great importance of this nutrient for maintaining health and well-being. However, food costs end up being characterized as not so favorable aspect.
This high cost is associated with the low availability of some ingredients used in animal feed, which results in high volatility and price increase. This means that food often corresponds to the largest share of expenses. As for example, in swine breeding, the costs of feeding can vary from 60 to 70%. In the case of fish production, animal feed represents 70% to 80% of total costs.
Therefore, the search for alternatives that are more economical has led to the research and discovery of new protein-rich ingredients that can be incorporated into animal feed, maintaining the same quality, in addition to contributing to increased productivity, performance, and development of the product.
Hydrolyzed proteins can be used to enrich the most varied food formulations, and it is a promising alternative to be applied in animal feed. And precisely for this application, the chicken protein hydrolysate stands out for being a highly palatable ingredient and for presenting a protein composition very favorable to the animal diet.
For all these characteristics and the extreme versatility, chicken protein hydrolysate is not limited to be used in the diet of only one animal species. On the contrary, it can be incorporated into the feed of pigs, dogs, cats, and fish.
Thus, it must be taken into account that each species has a specific need for amino acids and the protein content that they must consume daily. Therefore, to learn about hydrolyzed proteins and studies that relate them to animal performance is essential for formulators interested in using it as an ingredient in new diets.
In this blog post, we will deeply understand the benefits of using chicken protein hydrolysate in feed production and how this ingredient affects the performance and development of aquatic and land species.
Enzymatic hydrolysis: aspects to consider
Protein hydrolysis is aimed at breaking protein molecules, making them into smaller molecules, and providing desirable characteristics. This reaction can occur through chemical or enzymatic hydrolysis, in both ways changes in the functional properties and production of small peptides and amino acids will occur.
The hydrolysis of food proteins is performed for many reasons, including obtaining better nutritional characteristics, increasing the digestibility, delayed deterioration, conferring texture, increasing or reducing solubility, adding foaming or coagulation properties, adding emulsifying capacity, preventing interactions undesirable by removing off-flavors and toxic or inhibiting ingredients.
Hydrolysis can occur through the use of high temperatures (above 120 °C), however, this method causes the degradation of proteins, reducing the nutritional value of the food. Another way is through chemical hydrolysis, where the structures are modified through drastic changes in pH, either by acid or alkaline hydrolysis. This type of treatment is also aggressive and can lead to the degradation of proteins and amino acids, as well as the formation of protein complexes that decrease the digestibility of these nutrients.
Enzymatic hydrolysis is a reaction that occurs under mild conditions, both in temperature and pH, and pressure. In this way, the quality of the proteins, peptides, and amino acids generated, shows a great improvement in comparison to the other hydrolysis methods mentioned.
During enzymatic hydrolysis, proteolytic enzymes will promote the breakdown of proteins, generating smaller protein molecules, peptides, which have a molecular mass of less than 8000 Da, and some free amino acids. Thus, both peptides and amino acids favor the absorption of nutrients in the animal organism.
Some of these peptides, which are generally less than 3000 Da, are classified as bioactive, in which the specific sequence of their amino acids promotes biological actions that are beneficial to the animal, such as antioxidant, antimicrobial and anti-inflammatory activity, favoring the immune system and longevity.
The numerous advantages considering proteins hydrolyzed by the enzymatic process are definitely very attractive for those who want to produce high-quality ingredients. However, operating conditions must be strictly controlled to ensure the functionality of the final product. Some of these conditions that must be controlled are: type of enzyme used, enzyme concentration, substrate concentration, temperature, pH, pressure, agitation, in addition to activators and inhibitors of the reaction.
The type of enzyme used is a determining factor for the hydrolysis reaction, as each enzyme has a different specificity when it reaches its substrate. In this way, the enzymes have optimal conditions of activity (pH, temperature, pressure). When they are exposed to conditions that are not within their activity range, the enzymes change their molecular conformation, preventing the union between the substrate and the active site of the enzyme, thus not catalyzing the hydrolysis reaction.
Another important factor is the relation between substrate and enzyme. To a certain extent, the rate of enzymatic catalysis increases with increasing the amount of available substrate. However, after reaching saturation, that is, all the enzymes are already bound to the substrate, the reaction gets to a plateau and there is no more product formation.
However, enzymatic hydrolysis is the method considered most effective to obtain protein hydrolysates, as it presents high selectivity, there is no protein degradation, it generates products with bioactive peptides and with high bioavailability.
The importance of hydrolyzed proteins in animal nutrition
Proteins have numerous functions in the body of animals and are found in various cell structures, interstitial substances, antibodies, among others. Some important functions that can be attributed to proteins include: their role in the transport of oxygen (hemoglobin), in the protection of the body against pathogenic organisms (antibodies), as a catalyst for chemical reactions, membrane receptors, acting in muscle contraction (actin and myosin), in addition to being fundamental for the growth and formation of hormones.
However, some proteins have a complex structure, which reach the digestive system of animals and can’t be broken down and used for the biological functions more quickly and efficiently by the body. But peptides and amino acids are smaller molecules, and in general, they are fully absorbed in the digestive system of animals. In this sense, the hydrolysis of proteins, promotes this breakdown of complex molecules facilitating the digestibility and absorption of essential components for animals.
Considering this, it is essential to understand the structure of proteins and their complexity. Proteins are made up of peptides, and peptides are organic molecules formed by two or more amino acids linked together. The bonds between the amino acids occur between the carboxyl group of one molecule and the amino group of another and are called peptide bonds. In this way, peptides are differentiated according to the number and sequence of amino acids they present in their compositions. Peptides that have a molecular weight greater than 8000 Da are classified as proteins.
As for their structure, proteins are classified into:
- The primary structure of a protein corresponds only to its sequence of amino acids;
- Next, the secondary is the conformation of its polypeptide chains;
- The tertiary structure is the three-dimensional arrangement of the protein;
- Finally, the quaternary is the spatial arrangement of the polypeptide subunits.
The key to understanding the function of a protein is in its primary structure, which will define all other more complex structures and the biological activity of a given protein. In addition, the amino acid sequence has an influence on the functionality of peptides and proteins, as well as the presence of essential amino acids. Thus, it is clear that proteins are complex nutrients and linked to numerous functions in the animal organism, and so each species has different needs for amino acids as well as proteins.
Fish, being monogastric animals, do not have specific protein requirements. The main need in the diet of these animals is related to the essential amino acids. Thus, fish do not need large amounts of protein, but the proteins present in their food must be of high digestibility and present the essential amino acids.
What would be the ideal amino acid profile?
To be considered ideal, the profile must contain a total of 10 essential amino acids, not synthesized by the body. Therefore, these amino acids must necessarily be acquired through food. Cysteine and tyrosine are considered semi-indispensable, as there is the possibility of them being synthesized from methionine and phenylalanine.
The deficiency of essential amino acids reduces the growth and development of fish, which can develop anorexia. Besides that, this deficiency affects the immune system, resulting in animals more susceptible to infectious diseases.
In this sense, a study showed the deficiency of amino acids in the rainbow trout diet and its effects. Although it did not affect the production of antibodies, the activity of lysozyme and C-reactive proteins were reduced, affecting the defense mechanism of the animals.
Dogs & Cats
For land animals such as dogs and cats, there are some peculiarities regarding the requirement for amino acids.
Cats, being carnivorous animals, need a large amount of protein, preferably of animal origin. In general, it is recommended the consumption of 10 g of crude protein per day for young cats, 12.5 g for adults and 41 g for cats in the lactation phase. In addition, taurine and cysteine are essential amino acids for the diet of cats, which is different from the diet of dogs.
Considering dogs, it is recommended that puppies with a body mass of approximately 5 kg should consume up to 56 g of protein per day. In adult dogs with a body mass of 15 kg, it is recommended to consume 25 g of protein daily. Older dogs, on the other hand, need an even higher protein content, around 75 g of protein for 1000 kcal/day.
It is also important to consider the loss of amino acids that occurs in the gastrointestinal tract of dogs and cats. After digesting food, certain amino acids show greater losses in feces than others. This is the case, mainly, of aspartate, threonine, serine, and glutamate, which have the greatest losses for both species.
In order to compensate for such losses, the pets’ diet must be reinforced in these specific amino acids.
Swine also need the presence of 10 essential amino acids in their diets. In the case of these animals, it is necessary to pay attention to the limiting amino acids. These are those that, if not present, prevent protein synthesis from occurring. Lysine, methionine, threonine, and tryptophan are the limiting amino acids for swine, with lysine being the first one.
In addition, enzymatic hydrolysis can be a great ally in swine nutrition. Studies have already shown that the use of proteases to break proteins down into smaller, easily digestible peptides results in better pig growth and development.
A study on the Pig Progress website showed that the addition of a probiotic and a protease to the diet of more than 5,000 pigs led to an increase in the animals’ weight gain, in addition to higher survival rates (2% increase). Because of that, the use of hydrolyzed protein emerges as a viable option for swine nutrition.
Applications of chicken protein hydrolysate in animal nutrition
Chicken protein hydrolysate has a high nutritional value, developed with fresh, high-quality raw materials from poultry slaughter, such as offal and chicken meat, which is considered a functional protein specially formulated to improve the performance of animal feeds. It is produced through the enzymatic hydrolysis process, which generates smaller chains of amino acids and bioactive peptides.
This ingredient has all the essential amino acids, in addition to more than 70 different peptides having been identified and sequenced. Thus it is a suitable ingredient from the nutritional point of view for application in animal feeds, such as aquatic animals, pigs, dogs, and cats.
This improvement was confirmed mainly in aquatic animals, as the chicken protein hydrolysate proved to have a positive effect on the nutrition of aquatic species, such as tilapia, where it was found that 94% of this ingredient was digested. In addition, the availability of all amino acids present in the hydrolyzed protein was greater than 90%.
Tilapia also showed high attractiveness for diets containing chicken protein hydrolysate. The addition of 5% of the ingredient in the feed given to the animals increased the palatability index by 10.82% when compared to the control group, which contained 5% of fishmeal.
In terms of performance, tilapia had a 23% increase in weight gain with the inclusion of 2.5% chicken protein hydrolysate during the post-larval stage. The survival rate increased by 12% compared to the control group fed with fishmeal. The improvement caused by the chicken protein hydrolysate was also observed in other parameters, such as feed conversion rate (40% improvement) and final weight gain for juvenile fish (23% increase).
Chicken protein hydrolysate has also been tested in the diet of other species, with good results.
A study published in the Journal of Animal Science and Biotechnology demonstrated that the incorporation of this ingredient in a proportion of 6% in the feed of weaning pigs resulted in a significant increase in the growth of the animals, when compared to the control diet, without the addition of the hydrolyzed protein.
For dogs and cats, the palatability of chicken protein hydrolysate is especially attractive. In addition, the ingredient is highly digestible for these species, causing a considerable reduction in the production of feces. In the study by Revista Brasileira de Medicina Veterinária e Zootecnia, poultry by-products in dog food showed digestibility between 65 and 91%. It is worth mentioning that ingredients produced by enzymatic hydrolysis can increase these digestibility values.
Bioactive peptides and the strengthening of the immune system
Bioactive peptides are defined as fragments of amino acid sequences in a protein capable of conferring biological functions in addition to nutritional value. They generally have between 2 and 20 amino acids in the chain, a large abundance of hydrophobic amino acid residues and the presence of arginine, lysine and proline. One of the best known and most relevant actions for the animal organism is the antioxidant capacity that some bioactive peptides have.
Antioxidant activity is of great importance for the animal immune system, as oxidation is a natural process that occurs at all times in the body’s cells as a consequence of breathing. In general, oxidation leads to the formation of free radicals and reactive oxygen species that, to dissipate energy, react with other molecules in the cells, causing the process to spread. As a consequence, damage to lipids, carbohydrates, structural proteins, and DNA is caused. And these damages are related to a series of negative health problems, such as excessive inflammatory processes and the development of chronic and degenerative diseases.
To prevent oxidation from occurring in excess, cells also have antioxidant substances. These molecules react with free radicals and reactive oxygen species, stabilizing them and preventing them from reacting in a chain, damaging the macromolecules.
Thus, it can be said that the cells of the animal organism have a balance between pro-oxidant and antioxidant factors. When this balance is disturbed due to an excess of pro-oxidant factors, there is a condition known as oxidative stress. And if this condition remains active for a prolonged time, it can lead to the health problems mentioned here. In this sense, the consumption of bioactive peptides is essential to prevent these diseases.
Furthermore, in animals, bioactive peptides can assist in important physiological functions, such as antimicrobial activity and endocrine regulation. Once consumed, these peptides work in the small intestine, where there is the generation of signals transmitted to the brain, the endocrine system and the immune system, which has a positive impact on the entire body of the animal.
The real effects of bioactive peptides
A very interesting effect of bioactive peptides is the inhibition of the angiotensin-converting enzyme (ACE). ACE is involved in the formation of a vasoconstrictor peptide and is therefore associated with the development of hypertension.
Some bioactive peptides are able to inhibit ACE, reducing the risk of high blood pressure.
As antimicrobials, these peptides act by causing damage to the cell membrane of bacteria, interfering with the intracellular functions of their proteins, which induces the aggregation of cytoplasmic proteins, affecting the metabolism of the bacterium, which becomes unable to fulfill its functions.
The enzyme pepsin, under pH 5.5, the temperature of 23 ºC, a substrate concentration of 1% over 3 hours of reaction produces peptides of molecular mass between 445 and 2148 Da with antimicrobial and antihypertensive activity.
On the other hand, alcalase, pepsin, and trypsin enzymes produce peptides of 1000-2000 Da with antioxidant and anti-inflammatory activity if the hydrolysis is carried out at pH 7.0, at 50 ºC for 8h.
The presence of bioactive peptides in the chicken protein hydrolysate in the proportion of 2 to 3% in diets for tilapia (juvenile fish) increased the number of intestine villi in the animal’s organism. In addition, an increase in the nuclear area of hepatocytes has also been observed.
In adult tilapia, the inclusion of 2.4% chicken protein hydrolysate resulted in an increase of 6% when considering fillet yield. In these tests it was also observed that bioactive antiadipogenic peptides considerably reduced the content of total triglycerides and VLDL in animals, in addition to promoting an increase in levels of HDL, called good cholesterol.
Digestibility: an essential aspect to ensure protein absorption
The digestibility of proteins must be understood as being the part or portion of the protein that can be hydrolyzed by digestive enzymes into peptides and amino acids and, therefore, would be available biologically, as long as there was no interference in the capture of these nutrients by the animals’ organism. Protein digestibility is an important factor in determining the nutritional value of a protein, as it is directly associated with the absorption of these components.
Hydrolyzed proteins, in general, have a great advantage on protein digestibility, because as the molecules present in these products are smaller, it can be said that they are already partially digested, as it saves the hydrolysis of some enzymes that are in the digestive system. This way, these components are more easily absorbed by the animal organism, making digestion much faster and more efficient.
In a study, the chicken protein hydrolysate showed a high digestibility, in the in vitro system, obtaining values between 71 and 97%. This digestibility variation was due to the use of different enzymes in the hydrolysis process, and the hydrolysis of the chicken breast using the enzyme Alcalase was the one that presented the highest in vitro digestibility, being of 97%.
Tests carried out with chicken protein hydrolysate showed an in vivo digestibility of approximately 94% in tilapia and was proven to be considered an ingredient with a balanced amino acid profile for implementation in the feed of these fish.
The study carried out by Embrapa on the partial application of cassava protein in the feed of tilapia, suggests that vegetable proteins do not present a high digestibility for these animals, being more appropriate for the application of proteins of animal origin. The second group presents high digestibility and an increase in performance and quality of life.
Palatability: an essential characteristic to guarantee consumption of feed
We have already mentioned the quality of hydrolyzed proteins and the advances related to the availability of bioactive peptides in these ingredients. However, all these benefits would be in vain if the animals were not attracted enough to consume the feed with this type of protein source.
Palatability measures how desirable and attractive a given food is for animals. That is, when the animal comes into contact with the feed, how willing is it to consume it?
Pets, for example, have a much larger amount of aroma receptors compared to humans. This makes dogs and cats extremely sensitive to the smell, taste and texture of their food.
Thus, it doesn’t make sense that a formulation is developed for an animal if it is not palatable enough.
Some measurements can be done to determine the palatability of feed formulations:
- First choice (aroma): it measures, among a variety of foods, which is the food the most attracts the animal in the first contact. You can also measure how excited the animal becomes when approaching the analyzed food;
- Ingestion rate (flavour): this rate corresponds to the amount of the analyzed food consumed by the animal divided by the total amount of food consumed;
- Consumption rate (smell, texture, taste): it represents the preference that one population of animals shows for a formula compared to the others.
In the case of swine and aquaculture, poorly palatable feeds have a very negative effect on business profitability, resulting in animals with poor performance and development.
For swine, it is necessary to avoid protein sources that contain antinutritional factors, such as tannins, glucosinolates, and glycoalkaloids. It is also necessary to decrease the level of organic acids, responsible for a considerable drop in palatability.
For fish such as tilapia, some vegetable protein sources have already been reported in some in vivo studies to decrease palatability, negatively affecting the animals’ weight gain. This result was observed in studies with different parts of cassava added as ingredients in aqua feed.
On the other hand, hydrolyzed proteins produced from raw material of animal origin, such as chicken protein hydrolysate, have pleasant palatability and absence of anti-nutritional factors. These characteristics make, in addition to all the nutritional potential, the diets added with hydrolyzed protein to be attractive to animals, contributing to an improvement in performance.
Being fundamental parts of animal nutrition, in the most varied species, proteins and amino acids must be present in the diet in a balanced and biologically active way, always paying attention to the issue of essential and limiting amino acids.
In order to fulfill this requirement and produce beneficial effects on the development of animals, good protein sources need to be used as ingredients. In this scenario, hydrolyzed proteins, especially chicken protein hydrolysate, are produced by enzymatic hydrolysis from by-products of the agribusiness.
In addition to a high index of digestibility and palatability, the chicken protein hydrolysate can present bioactive peptides that fulfill essential functions in the animal organism, keeping them healthy and free from problems related to growth and the immune system. As well as adding value to industry co-products, reducing the possibility of waste and environmental risk.