Date: 06/04/2020

According to the Food and Agriculture Organization of the United Nations (FAO), global fish supply is projected to reach 201 million tonnes in 2030. Production from aquaculture is expected to hit 109 million tonnes, representing a 36% increase from the 2016 production of 80 million tonnes.

Aquaculture production still relies heavily on fish meal and its supply chain faces many challenges. First, we know that the demand for fish meal is growing faster than the available supply, leading to price increases. In terms of costs, in 1983, the average fish meal price was around USD400/tonne but in recent years it has gone up to USD1,600/tonne. As a comparison, soybean meal price increased eight times less in the same period.

Other than increasing prices, there is also the issue of availability of high-quality fish meal, which are produced with whole fish and contain less ash content. The supply of such fish meal (~68% crude protein), is getting less. This is because of lower catches from capture fisheries which in turn limit the fish meal supply for aquaculture. In addition, the use of this raw material is considered unsustainable in the medium or long term.

As our source of fish protein evolves from capture fisheries to aquaculture, it also means that we need to provide farmed fish with all the required nutrients in a complete and balanced feed.

Furthermore, each time we move further to more intensive productions, there is less dependence on natural feeding. The quality of ingredients used in feed formulations and the type of feed processing used have a direct impact on fish productivity.

Different processes for producing ingredients and their characteristics 

To understand these differences, this article will provide some explanations on three types of ingredient processing; the most common is thermal processing or cooking. This happens when the raw material is subjected to high temperatures in a digester. This process has the benefit of being simple and cheap, but it may be detrimental to the proteins and impacts the digestibility and availability of amino acids. For this process, studies indicate that the longer the raw material is exposed to high temperatures, the lower the digestibility and nutritional value.

The second process is chemical hydrolysis, which can be acid or alkaline. This is when the raw material is subjected to severe pH changes. This type of hydrolysis is also considered a low-cost process. Acid hydrolysis is often used to enhance flavour but a negative point is that with this process, there will be a partial destruction of some amino acids, such as tryptophan.

When we talk about alkaline hydrolysis, the impact on amino acids is even worse than of acid treatment. Another fact is that chemical hydrolysis will increase the ash content in the ingredient, since you are adding other components during the process.

Peptide bond enzymatic hydrolysis scheme.

Figure 1. Peptide bond enzymatic hydrolysis scheme.

Lastly, we have the enzymatic hydrolysis, which often happens at low temperature and low pressure and does not result in any loss of amino acids. Furthermore, it improves the digestibility of protein and availability of amino acids to the animal.

Enzymes are also more precise in controlling the degree of peptide-bond hydrolysis, making it possible for a greater consistency of the final product. Another advantage is that in this process, we have less use of energy and steam compared to the cooking process.

Enzymatic hydrolysis and bioactive peptides

To understand more on how enzymatic hydrolysis generates high quality ingredients with functional benefits, we will review some concepts. By definition, a protein is a macromolecule that usually has 20 different amino acids (AAs) that are linked via peptide bonds. Amino acid contains both amino and acid groups. The peptide bond is the link between the nitrogen from the amine group with the carbon from the acid group. The sequence of AA is what defines the different proteins and its biological value.

When we have a protein with a low molecular mass, and a low number of amino acids, we call it a peptide. Another important fact is that peptides may have different bioactivities such as antimicrobial, antioxidant, antihypertensive and also immunomodulatory.

These are called bioactive peptides, and are defined as specific sequences of amino acids, with low molecular mass and a specific biological activity in the organism.

One way to produce bioactive peptides is through enzymatic hydrolysis of the intact protein. The word “hydrolysis” means breaking through the water.

The enzymatic hydrolysis is a chemical reaction accelerated by an enzyme that uses water to break one molecule into two other molecules. In the case of proteins, the hydrolysis forms smaller chains of amino acids (peptides). These small peptides are easily absorbed into the animal’s organs than a whole protein. In addition, the animal will spend less energy absorbing them. This extra energy can be used to grow, to gain weight and to fight challenges.

The protein digestive capacity depends on several factors, among which are the age of the animal (i.e. larval stage or adult) and the protein form (if it is a small peptide, a free amino acid or an intact protein). Usually, more mature animals will absorb food more efficiently than animals in the early stages; thus, hydrolysed proteins will be more easily absorbed than intact proteins. Besides the benefit of in vivo protein absorption, enzymatic hydrolysis also generates bioactive peptides.

There are extensive experimental stages involving enzymatic engineering knowledge to determine which is the best enzyme for which raw material, and what processing conditions are needed to achieve the desired bioactivities. The bioactivity can vary depending on the type of enzyme used and each enzyme can create a peptide with a different AA sequence.

Different types of raw materials have different specific peptides and bioactivities, such as immunomodulatory and antimicrobial. Those bioactive peptides identified can be further explored in the future for benefits in animal nutrition, opening a wide variety of applications for protein hydrolysates.

Protein hydrolysates as functional ingredients

There is a clear relationship between the way how ingredients are processed (and hence their nutritional value) and the animal performance. With that in mind, BRF Ingredients has designed a new method of processing ingredients based on enzymatic hydrolysis; this process has given better results for animal performance.

Molecular mass distribution of BRFi hydrolysate product.

Figure 2. Molecular mass distribution of BRFi hydrolysate product.

After enzymatic hydrolysis of the raw material, almost all the solids are removed through filtration, thus decreasing the final ash content to approximately 4%, and concentrating the protein content to more than 75%, with a digestibility of more than 90%. The stability of these parameters is another characteristic of the process. Furthermore, with the selection of enzymes and process conditions most of the protein content is in the form of small peptides with less than 3 kDa of molecular mass, that is considered to be the band with greater bioactivity.

In addition to the advantages of the process itself and the characteristics of the final ingredients, BRF as a 100% integrated company can easily guarantee the traceability and freshness of the raw material used.

Impact on animal performance

In in vivo trials with tilapia, the protein digestibility was 93.61% and the inclusion of 2.5% of chicken protein hydrolysate in formulations of tilapia in initial stages increased the final weight by 23% in comparison with the control group with 10% fish meal. The experiments also showed a gut health promoter effect with the inclusion of 3% of protein hydrolysate, which increases the number of villi by 49% compared to fish meal control group.

In adult tilapia, the inclusion of 2.7% protein hydrolysate in their feed increased the fillet yield by 6% compared to the fish meal control group. Besides that, the analysis of the plasma lipid profile of the tilapia showed that, with the inclusion of protein hydrolysate, the levels of triglycerides and VLDL (very low density lipoproteins) decreased, while levels of HDL (high density lipoproteins) increased significantly (Figure 3)

Plasma lipid profile of tilapia fed with diets containing different levels of inclusion of protein hydrolysate ingredient(experiment performed at the Western Paraná State University-Unioeste, Brazil).

Figure 3. Plasma lipid profile of tilapia fed with diets containing different levels of inclusion of protein hydrolysate ingredient(experiment performed at the Western Paraná State University-Unioeste, Brazil).

This result is an indication of the presence of anti-adipogenic bioactive peptides, which improved the energetic metabolism and promoted a rational utilisation of nutrients in protein deposition.

In a study developed in Vietnam, 2% of protein hydrolysate was shown to be able to replace 5% of fish meal and was able to maintain growth performance of the tilapia. In this case, besides the benefit of bioactive functionalities, the change from fish meal to protein hydrolysate has the advantages of ingredient restorability, price and quality parameters stability, with no significant effect in formulation cost.

With regards to results on the application of protein hydrolysates in shrimp feed, enzymatic hydrolysate ingredients yielded 94% of digestibility coefficient of the protein for this specie. In growth experiments, the inclusion of 5% of hydrolysate proteins in the formulation improved the final weight by 7% and the feed conversion ratio (FCR) by 14.3% compared to the salmon meal control group.

Cumulative mortality of Litopenaeus vannamei fed with diets containing different levels of hydrolysate protein after 48h post-infection (h.p.i) with Vibrio parahaemolyticus at the concentration of 9 x 107 CFU/mL (experiment performed at Universidade Federal de Santa Catarina, Brazil (UFSC).

Figure 4. Cumulative mortality of Litopenaeus vannamei fed with diets containing different levels of hydrolysate protein after 48h post-infection (h.p.i) with Vibrio parahaemolyticus at the concentration of 9 x 107 CFU/mL (experiment performed at Universidade Federal de Santa Catarina, Brazil (UFSC).

In a challenge test to evaluate the resistance of shrimp intentionally infected with a Vibrio bacterium, all the treatments that had inclusions of protein hydrolysate ingredients in the formulation showed a reduction between 20% and 50% in the cumulative mortality compared to the control group which contained only fish meal (Figure 4). This is a strong indicator of the immunomodulatory and/or antimicrobial effect of bioactive peptides.

In aquaculture, the short supply of some important ingredients can act as a catalyst for the development of smarter alternatives bringing better value to the business. There are still many secrets on hydrolysate proteins and bioactive peptides, but there are already many studies reporting benefits in medicine, cosmetics, human nutrition and also in animal nutrition.

In animal nutrition specifically, results from the application of peptide products and hydrolysed proteins in diets of different species have shown benefits including improved intestinal health, immune system, growth and production performance.

In an increasingly competitive market, these advantages should not be neglected. The whole world is changing and evolving rapidly and in the area of ingredients for animal nutrition it is not different. New technologies and smarter processes arise all the time and it is up to us to enjoy the benefits and contribute to a more efficient and productive chain and business.

Authors: Thaís Costa Andrade is R&D Specialist and Wilson Rogério Boscolo is R&D Consultant at BRF Ingredients Brazil.