INTRODUCTION
In animal rearing, feeding is one of the most important and expensive activities. This fact has a special relevance, considering the increase in recent years of the price of raw materials used for this purpose. For these reasons, the search for alternatives that allow the inclusion of new by-products, as well as the efficient use of the nutritional value of foods, represent a priority for animal science.
The use of zootechnical additives is one of the strategies used for improving productive performance of animals and reduce production costs (Carro et al. 2006). This category includes probiotics, prebiotics, plant extracts and enzymatic preparations. The use of this last group in conventional and alternative diets in different species allows to improve productive indicators and health of the supplemented animals (Ferreira et al. 2016 and Yamabhai et al. 2016).
This objective of this paper was to provide updated scientific information on obtaining hydrolytic enzymes and their use in the feeding of monogastric species.
ENZYMATIC ADDITIVES
Enzymes are proteins with catalytic activity that possess extraordinary efficiency and specificity, and its function is to catalyze the chemical reactions that occur in living cells (Garg 2016). The increase of research related to enzymatic technology allows the applications of these proteins to be extended to different fields such as industries producing ethanol, detergents, paper, and some other products, as well as in animal feed.
Exogenous enzymes are considered to be those that do not belong to the digestive system of animals, so they must be included into the diets (Rojo et al. 2007). The use of these proteins in animal feed was first reported in 1925 (Bedford 2018). During the 50s, studies were conducted to evaluate their addition to rations for poultry. Results were variable and, generally, with little productive response (Brenes 1992).
In the 80s, the use of these additives in animal feed began. Pioneer countries in its application were the Scandinavians, Great Britain, Canada and, in a very particular way, Spain (Brufau 2014). These proteins were initially used in wheat and barley producing countries, currently their application extends to other grains, such as corn, soybean and sorghum. The benefits of using this technology in animal diets are associated with improved yield and reduced rearing costs (Souza et al. 2014).
In recent decades, the use of enzymes has expanded, so that 200 million tons of pig and poultry feed are supplemented with these products (Graham and Bedford 2007). For 2019, it is estimated that the market value of enzymes for animal production will reach 1,280 million dollars, with a growth rate of 7.6% for the period between 2014-2019 (Brufau 2014).
Enzymatic additives can exert their effects through direct actions on food, before they are consumed or from modifications of digestive processes of animals in which they are applied (Carro et al. 2006). Generally, its use is aimed at solving two fundamental problems: improving availability of polysaccharides, lipids and proteins, which are protected from digestive enzymes by impermeable structures of the cell wall of plants, and degrading materials that interfere with digestion, absorption and utilization of nutrients (McDonald et al. 2010).
Its application facilitates the best use of nutrients in feed and makes possible the use of lower quality ingredients, alternative raw materials and by-products. They are also used in order to improve total digestibility of diets, increase digestibility of certain nutrients, complement the activity of endogenous enzymes produced by the animal, especially in young poultry and pigs, as well as reduce the excretion of certain compounds such as phosphorus and nitrogen (Cortés et al. 2002, Carro et al. 2006 and Fernández and González 2011).
This analysis should take into account that digestive processes in animals are not completely efficient. Pigs and poultry cannot digest 15-25% of the food they consume, because it contains indigestible antinutritional factors that interfere with digestive processes. In addition, animals do not have specific enzymes that break certain food components (Bedford and Partridge 2010).
ANTINUTRITIONAL FACTORS
Non-starch polysaccharides (NSP). NSPs are the main components of plant cell walls. These include cellulose, hemicelluloses, pectins, β-glucans and other polysaccharides (Gray 2006). Arabinoxylans are the major NSPs in wheat, rye and triticale, while β-glucans are the most abundant in barley and oats. In other feed ingredients, such as soybean, sunflower or rapeseed, other types of NSPs like β-galactosides and β-galactomannans appear in lesser amounts, but with a marked anti-nutritional effect (Fernández and González 2011).
NSPs are classified as soluble and insoluble (Lata 2011). These compounds constitute a barrier to the action of hydrolytic enzymes, by retaining the rest of the nutrients in the endosperm cells (Willians et al. 1997). In addition, its presence is related to an increase in the viscosity of the digesta, lower digestibility of fats, proteins and carbohydrates and a decrease in the activity of endogenous enzymes, resulting in reduced contact between these proteins and nutrients (Nikam et al. 2017).
Excess of these compounds in the diet can cause changes in the microflora of the gastrointestinal tract. In poultry fed with high contents of these macromolecules, a large amount of anaerobic bacteria was observed, which may be associated to increased intestinal viscosity (Dudley-Cash 2014).
The inclusion of foods rich in NSP in diets for broilers may cause lower yields in animal production parameters (Alba 2013). In the case of pigs, it is known that the use of diets with wheat by-products and high levels of these polysaccharides, reduces nutrient digestibility (Nortey et al. 2007).
García (2000) analyzed the effectiveness of several applied methods, in order to increase the nutritional value of cereals rich in NSP. This author recommended, as the most widespread possibility in practice due to its effectiveness and ease, the addition of enzymes of microbial origin with β-glucanase and xylanic activity, which allow the rupture of β-glucans and xylanes, respectively, and avoid the inconveniences derived from the presence of these compounds.
Phytic acid and phytates. Phytic acid is the main form for storing phosphorus in plants. It is widely distributed in cereals, legume grains and seeds. It is also found in pollen, roots, stems and leaves (Shanmugam 2018). It represents more than 70% of the phosphorus content in cereals, mainly in corn and wheat, which are essential raw materials for animal feeding. Because phytic acid is unstable in its acid form, it is mainly found as phytates. In this way, it forms compounds associated with divalent metal cations and with some proteins and amino acids (Humer et al. 2015). Phytates are anti-nutritional factors because they are insoluble salts, negatively charged, limit phosphorus, mineral and protein availability, which cannot be absorbed in the gastrointestinal tract of monogastrics due to the low levels of endogenous phytases (Chen et al. 2015 and Ingelmann et al. 2019).
Soto (2015) considers that phytates can reduce the activation of the stomach pepsin enzyme, responsible for the digestion of proteins, and can negatively affect the mechanism by which amino acids are absorbed from the small intestine.
The application of phytase enzymes in the diet allows to eliminate the anti-nutritional effects of phytates and increases the productive results of poultry. Given that enzymes are natural, biodegradable, and non-toxic products, their use in the nutrition of monogastric species is one of the most promising alternatives for reducing the negative effects of anti-nutritional factors in animal rearing.
SOURCES FOR OBTAINING ENZYMES
Most of the enzymes used in food and food processing have a microbial origin. These proteins are generally produced from bacteria (Bacillus subtilis, Bacillus lentus, Bacillus amyloliquefaciens and Bacillus stearothermophilus), fungi (Triochoderma longibrachiatum, Asperigillus oryzae and Asperigillus niger) and yeasts, such as Sacchoromyces cerevisiae (Khattak et al. 2006). Filamentous fungi, specifically Aspergillus and Trichoderma genera, have been widely used, due to their ability to secrete xylanolytic enzymes to the culture medium (Fadel et al. 2014).
Other authors recommend the use of bacterial cultures for these purposes, and point out that the low nutrient demand of these microorganisms for their growth and their possibilities of development in varied environments are among the most important factors to take into account for this election. These authors also emphasize their ability to produce stable enzymes under extreme conditions of temperature and pH, which could be maintained in bioconversion processes, as well as to increase the rates of enzymatic activity, fermentation and product recovery (Maki et al. 2009 and Chakdar et al. 2016).
Bacillus is one of the most used genera for this purpose, which is distributed in various habitats, and can survive under adverse conditions, due to their endospores production (Kim et al. 2017). Specifically, Bacillus subtilis is an abundant and stable bacterium that is not considered pathogenic and shows potential characteristics for its use in the preparation of zootechnical additives (Milián et al. 2014, Cheng et al. 2016 and Milián et al. 2017). It has an excellent fermentation capacity and offers the advantage of secreting great amounts of enzymes into the culture medium (Meima et al. 2004), such as proteases (Akcan and Uyar 2011), amylases (Maity et al. 2015), mananasas (Pangsri and Pangsri 2017) and xylanases (Sugumaran et al. 2013 and Ho 2015).
Current processes for enzyme production are based on the use of genetically modified microorganisms. This methodology increases the productive capacity of the fermentation unit itself and avoids the presence of undesirable activities (Brufau 2014 and Garg 2016).
Most of the marketed industrial enzymes are produced by submerged fermentation. However, solid state fermentation improves yields, facilitates the separation of the obtained products and lowers production costs, by using agricultural waste materials to obtain the desired enzymes (Zhang et al. 2012 and Fadel et al. 2014).
The use of these by-products helps to solve one of the main problems of the current society, which is the generation of large volumes of waste, with the consequent risk to the environment and public health (Serrano 2015). Table 1 shows examples of the production of enzymes with potential for application in animal feed through the use of solid state fermentation and the use of different by-products.
Enzymes | Microorganisms | Substrata | Author |
---|---|---|---|
Lignin peroxidase and β-Glycosidase |
|
Sawdust | (Kamsani |
Xylanase | Wheat bran | ( Ho |
|
Xylanase | Corn | (Helianti |
|
Xylanase | Corn meal | ( Kallel |
|
Xylanase and β-mannanase | Mixture of corn and wheat bran residues | (Zhang and Qing, 2015) | |
β- Mannanase | Torta de harina de palma | (Siti-Noritaac |
|
Xylanase | Salvado de trigo | (Phadke and Momin 2015) | |
Proteases | Afrecho de trigo | (Pouryafar |
FACTORS THAT INFLUENCE ON EFFECTIVENESS OF ENZYMATIC PREPARATIONS
Effectiveness of enzymatic supplementation in animal nutrition depends on various factors related to the characteristics of enzymes and of the animals in which they are applied. The source from which these proteins are obtained, applied dose, collateral activities of other enzymes present, diet composition, age, health and productivity of supplemented animals can limit the success of the used treatment (Bedford and Cowieson 2012 and Salem et al. 2013). The influence of some of these factors is briefly explained below.
Animal species. Enzymatic additives are used, to a greater extent, in monogastric species, mainly in pigs and poultry. The characteristics of poultry gastrointestinal tract contribute to a more uniform response, compared to pigs. The presence of the crop favors the influence of enzymes on the substrates present in the feed, before reaching the stomach. During the period of food storage in this organ, pH is 6.3, favorable value for the performance of most of these enzymes. In pigs, food is retained for a long period in the stomach, with more acidic pH values than in poultry, which could cause the inactivation of the used enzymes. Another aspect to consider is that poultry have less capacity to ferment fiber than pigs (Graham et al. 1988 and McDonald et al. 2010).
In ruminants, the use of enzymes in their diets was limited in the past, since it was assumed that they can be inactivated during rumen proteolytic activity. However, during the past decade, it was demonstrated that enzyme preparations can be effective for improving lactation and growth in bovine cattle (Plumstead 2013).
Age of animals. Exogenous enzyme supplementation with food is more successful in young animals. These categories are characterized by having lower digestive capacity and, in general, their enzyme system is not fully developed. Studies in pigs and poultry showed that the benefits of enzymatic supplementation of diets decrease with age increase (Ravindran 2010).
Diet composition and quality. Proper use of enzymes in animal feeding requires a careful selection of diet ingredients to obtain economic benefits (Asmare 2014). Animal response depends on the quality of these ingredients. The lower the quality, the greater the magnitude of the improvements obtained by enzymatic supplementation (Ravindran 2010). In this case, diet quality largely depends on its content of anti-nutritional factors. The effectiveness is also related to the used cereal, cultivar and level (Brenes 1992). This fact is related to the specificity of enzymes and their affinity for certain substrates.
The selection of the enzyme preparation to be used should be made when considering the nature of the substrate that will be degraded (Brenes 1992). For example, wheat contains arabinoxylans that limit the digestibility of its nutrients. It was experimentally demonstrated that this difficulty can decrease with the addition of xylanases in the diet of poultry fed with this cereal (Nortey et al. 2007).
It should also be considered that various ingredients with different chemical structures are used in the diet for monogastric species, so it is recommended to use a combination of several enzymes to achieve better results (Duran 1992 and Dudley-Cash 2014).
EFFECTS OF ENZYMATIC ADDITIVES ON ANIMAL HEALTH AND PRODUCTIVITY
The inclusion of phytases, carbohydrases and proteases in monogastric diets has generated great interest in recent years. Supplementation with these enzymes had a positive impact on productive indicators of these animal species (Asmare 2014).
Phytases can degrade phytate from plants that are used as ingredients in animal feed (Graham and Bedford 2007). Its use in poultry and pigs caused increases in phosphorus availability, between 20 and 45% (Ravindran 2010). Energy utilization and amino acid availability in the diet increased, which improved indicators such as conversion and production of meat and eggs (Bedford and Partridge 2010). The inclusion of these enzymes in poultry diets helps preserve the environment and prevent contamination by decreasing phosphorus excretion in feces (Selle and Ravindran 2007).
The β-mannanase enzymes hydrolyze the mannane into oligosaccharides, compounds that have prebiotic interest (Yamabhai et al. 2016 and Zuluaga et al. 2017). Its effects as promoters of animal health and growth were verified in different animal species, such as turkeys, poultry and bovines (Yamabhai et al. 2016 and Seo et al. 2016).
The addition of this enzyme to poultry diets caused a decrease of food intake, which can be attributed to the improvement of nutrient absorption related to the decrease of digesta viscosity and to the increase of villi height in the duodenum. This last factor indicates that nutrient absorption was superior in this organ. It was also found that supplementation with this protein improved weight gain and food conversion (Imran et al. 2014). In addition, Rehman et al. (2016) recommend valuing their utilization in this species, when low energy diets are used.
The application of enzymatic additives also causes changes in the intestinal microbiota, which can have a beneficial impact on the health of animals that consume them. Some of the mechanisms involved are the increase of nutrients released from the host, formation of fermentable oligosaccharides, as a result of the depolymerization of insoluble fiber, and the acceleration of digestion that produces rapid movement of fermentable carbohydrates and proteins (Bedford and Cowieson 2012).
In the case of ruminants, enzymatic additives have been used to improve food use efficiency and to reduce waste production in animal production systems (González 2004). The introduction of these proteins is one of the strategies used for increasing starch digestion rate and the energy value of grains in these animal species (Plata et al. 2004).
The addition of xylanase to polygastric diets improves the digestion of food derived from plants and produces compounds with nutritional value for ruminal flora (Garg et al. 2010). The application of this enzyme to forages, combined with cellulases, increased silage quality and, consequently, the range of digestion of the cell wall by ruminants (Garg 2016).
Mixtures of enzymes, such as amylases and cellulases, which improve the digestibility of plant cell walls and/or starch, can increase cattle productivity (Rojo et al. 2001 and Gutiérrez et al. 2005). Other effects of enzyme supplementation in different animal species are presented in Table 2.
Enzymes | Species | Effect | Author |
---|---|---|---|
β-mannanase | Poultry | Increase of the daily mean gain and improvement of conversion | (Ferreira |
β-mannanase y β-glucanase | Poultry | Favorable morphological changes in the small intestine | (Karimi and Zhandi 2015) |
Cellulase and xylanase | Goats | Increase of milk production and weight gain | (Trejo |
Enzymatic additive with xylanase, glucanase and mannanase activity | Poultry | Health improvement, attenuation of growth retardation of poultry, challenged with |
(Sun |
β-mannanase | Red tilapia | Growth improvements | (Siti-Noritaac |
β-mannanase | Bovines | Increase of growth and feed efficiency | (Seo |
β-mannanase | Growing pigs | Reduction of the number of fecal coliforms and tendency to decrease the emission of ammonia from feces. | (Upadhaya |
Combination of xylanase, ß-D-glucanases, cellulases, mannanases and peptinases | Poultry | Increase of yield, better food intake and food conversion improvement | (Nikam |
The use of enzymatic additives in animal nutrition is considered to be a promising option from an economic, environmental and sustainability point of view (Asmare 2014). In addition, this practice offers new opportunities in the market for crops such as canola, sunflower and cotton, because they favor the use of nutritional properties of these alternative sources (Souza et al. 2014).
GENERAL CONSIDERATIONS
Obtaining enzymes from microorganisms, with the use of agroindustrial by-products as substrates, is a strategy that reduces the production costs of these additives. In addition, it encourages the proper use of waste materials that contribute, many times, to environmental pollution.
Enzymatic additives are used in the nutrition of monogastrics, especially in poultry and pigs. Its use allows to increase the use of raw materials, in many cases of low cost, in the supplied diets.
Although several researches support the positive results of the application of these additives in increasing nutrient digestibility and improving the productive indicators of supplemented animals, there is great variability in them. This may be related to the diversity of factors involved in its effectiveness, among which those of these enzymes stand out (their stability, specificity of action and some others), those related to the animals in which they are used (species, age and morphophysiology of the gastrointestinal tract) and the characteristics of the supplemented diets.