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Enzyme Hydrolysis Products and Yeast-derived Products as Prebiotics and Natural Alternatives to Antibiotic Growth Promoters

Published: January 27, 2014
By: Bogdan A. Slominski (Department of Animal Science, University of Manitoba)
The objective of this presentation is to discuss the role of dietary enzymes in the production of hydrolysis products that are important for animal health and food safety in the antibiotic-free poultry and swine nutrition. This presentation would also address any potential relationship between feed enzymes and yeast-derived products with the emphasis on the development of new enzyme supplements effective in improving the functionality of yeast-containing feed ingredients, including distillers dried grains with solubles (DDGS). 
Prebiotic effect of NSP-hydrolysis products
Carbohydrase enzymes have a direct, positive effect on animal performance by improving nutrient digestion and absorption, thereby reducing substrate availability for microbial growth in the ileum (Choct et al. 1999; Bedford and Apajalahti 2001, Slominski 2011). In the process of depolymerizing various polysaccharides (NSP) in the diet, carbohydrase enzymes may produce galacto-, gluco-, manno-, or xylo-oligomers (Silva et al. 1983), which, similarly to prebiotics, may facilitate proliferation of bacteria beneficial for gut health such as Bifidobacterium and Lactobacillus, thereby decreasing the abundance of pathogens such as Clostridium, Salmonella, E. coli and Campylobacter (Gibson and Roberfroid 1995). In this context, these enzyme hydrolysis products may indirectly prohibit the growth of certain pathogenic species by increasing acidity through an increase in lactic acid in the lower gut. Concomitantly, non-substrate utilizers, in a highly competitive ecosystem, will be suppressed and can virtually disappear. In this context, the use of lactic acid bacterial cultures Lactobacillus acidophilus and Streptococcus faecalis has shown promising results in suppressing C. perfringens proliferation (Fukata et al. 1991) and reducing C. perfringens associated mortality (Hofacre et al. 2003). In addition, certain enzyme hydrolysis products may attract microbes away from the intestinal binding sites by a means of competitive exclusion, thereby reducing colonization and disease and allowing the mucosa to perform its function of secretion, digestion and nutrient absorption.
Commonly used feedstuffs such as wheat, corn, barley, soybean meal, canola meal, peas, wheat by-products, and flaxseed contain significant amounts of non-starch polysaccharides (NSP), including rhamnogalacturonans, galactomannans, mannans, arabinoxylans, β-glucans, arabinans, arabinogalactans, etc. As illustrated in Table 1, incubation of soybean meal, canola meal and flaxseed meal with a multicarbohydrase enzyme resulted in reduced amounts of water-insoluble NSP and increased amounts of water-soluble NSP and NSP hydrolysis products with some monosaccharides being released in the enzyme-treated samples. Among the monosaccharides released, glucose, galactose and uronic acid were predominant in all meal samples (Jia et al., 2009). It would appear that when NSP are broken down into low-molecular weight polysaccharides, oligosaccharides and simple sugars by the correct blend of carbohydrase enzymes they acquire the potential to become prebiotics and can in turn exert health benefits by improving the intestinal environment.
Recent two studies conducted in our laboratory demonstrated that the addition of a newly developed multi-enzyme preparation containing pectinase, cellulase, mannanase, galactanase, galactosidase, xylanase, glucanase, and other enzyme activities was effective in promoting growth and feed utilization of broiler chickens fed an antibiotic-free diet and facilitated post-disease compensatory growth of broiler chickens challenged with C. perfringens, a causative agent of necrotic enteritis (Table 2). In this study enzyme supplementation was accompanied by a 1.3 log reduction in C. perfringens counts (from 4.3 to 3.0 log10 CFU/g). 
Table 1. Nonstarch polysaccharide (NSP) and NSP hydrolysis product balance after incubation of ethanol-extracted soybean meal, canola meal and flaxseed meal with a multicatobohydrase enzyme (mg/g DM).
Enzyme Hydrolysis Products and Yeast-derived Products as Prebiotics and Natural Alternatives to Antibiotic Growth Promoters - Image 1
Table 2. The effects of diet, enzyme addition and C. perfringens challenge on growth performance of broiler chickens.
Enzyme Hydrolysis Products and Yeast-derived Products as Prebiotics and Natural Alternatives to Antibiotic Growth Promoters - Image 2
When using an advanced “in situ” experimental model, the infusion of NSP hydrolysis products into living piglet intestinal segments that were experimentally infected with E. coli K88 was investigated and the fluid passage and absorption (to estimate diarrhea) through the segments were measured (Kiarie et al., 2008). It was found that after an E. coli K88 infection, segments that were infused with enzyme hydrolysis products had greater fluid absorption than control segments (Figure 1). This means that the sugars released following NSP hydrolysis exert a beneficial effect on intestinal tissues during an infection which could lead to reduced scours and improved recovery in a commercial operation.
In a similar study from this laboratory (Kiarie et al., 2009), piglets challenged with E. coli K88 and provided with NSP hydrolysis products from canola meal, soybean meal, and flax had lower stomach pH, reduced intestinal fluid loss and greater levels of lactic and total organic acids compared to control piglets (Table 3). In addition, piglets fed enzyme hydrolysis products consumed more feed, grew better and had lower incidence of diarrhea than control piglets. In this study NSP hydrolysis product addition was accompanied by the reduction in total coliform and E. coli K88 counts and a significant increase in beneficial Lactobacilli counts (from 6.9 to 9.4 log10 CFU/g). 
Table 3. The effect of NSP hydrolysis products on growth performance and the intestinal environment of piglets 24-hours after E. coli K88 challenge.
Enzyme Hydrolysis Products and Yeast-derived Products as Prebiotics and Natural Alternatives to Antibiotic Growth Promoters - Image 3
Figure 1. Development of diarrhea (measured as fecal scores) of piglets fed NSP hydrolysis products and challenged with E. coli K88. Higher fecal scores indicate more severe diarrhea.
Enzyme Hydrolysis Products and Yeast-derived Products as Prebiotics and Natural Alternatives to Antibiotic Growth Promoters - Image 4
It is evident from this research that the benefits to be gained from enzyme supplementation are not only from improved nutrient digestion and feed efficiency. Improved gut health as a result of prebiotics formed from the hydrolysis of common feedstuffs can also benefit the feed industry by controlling enteric infections. 
Yeast-derived products and yeast cell wall lytic enzymes
Yeast products are rich sources of mannan polysaccharides, ß1,3- and ß1,6-glucans, and nucleotides, which can function as prebiotics and have been shown to stimulate the immune system and gastrointestinal tract development (Zhang et al. 2005; Zdunczyk et al., 2005; Solis de los Santos et al., 2005) and to provide favorable conditions for beneficial intestinal Lactobacillus spp., and competitive binding sites for pathogens with mannose-specific fimbriae such as Salmonella, thus decreasing attachment and colonization. In addition, yeast cell wall ß1,3- and ß1,6-glucans have been reported to provide protection from the deleterious effects of E. coli challenge in broiler chickens (Huff et al., 2006). Little is known about yeast nucleotides as they are not considered essential nutrients. Some research during the last several years indicated that this may not be completely correct since under periods of rapid growth and metabolic stress, demand may exceed the capacity of de novo synthesis and exogenous dietary nucleotide supplementation may spare the energetic cost of de novo synthesis.
The potential of a yeast product rich in nucleotides to reduce medication use in broiler chicken diets was evaluated using 1600 male Cobb x Cobb broiler chickens, placed at 1 day of age, in a completely randomized experimental design. The test article used in this study was Maxi-Gen PlusTM, a nucleotide-rich yeast product containing a mixture of mono-nucleotides. Birds were randomly assigned to the following dietary treatments: Positive control (PC) containing 110 ppm bacitracin methylene disalicylate (BMD), Negative control (NC; no medication), NC + 0.05% yeast nucleotides (YN), and NC + 0.05% YN + 55 ppm BMD. Diets were fed ad libitum for 42 d.
As illustrated in Table 4, birds fed a diet containing YN + BMD had the same BWG as PC birds (2.35 vs. 2.33 kg) and both treatments had greater BWG than NC (2.20 kg) and NC + YN (2.28 kg) birds. However, BWG of NC + YN birds was significantly higher than that of NC. Feed intake was significantly greater in YN (4.23 kg/bird) and YN + BMD birds (4.25 kg/bird) than in NC birds (4.09 kg/bird) and did not differ (4.20 kg/bird) compared to PC birds at d 42. There was no difference in FCR between YN + BMD and PC birds (1.81 vs. 1.83) and YN + BMD birds had lower FCR than both NC and NC + YN birds. Mortality was lower in YN birds than NC birds (1.8% vs. 5.3%) and did not differ compared to YN + BMD or PC birds. This study suggests that feeding broiler chickens a yeast product rich in nucleotides can assist in reducing dietary medication usage. 
Table 4. The effect of yeast nucleotides on growth performance of broiler chickens (1-42 d).
Enzyme Hydrolysis Products and Yeast-derived Products as Prebiotics and Natural Alternatives to Antibiotic Growth Promoters - Image 5 
Over the last few years, the author of this article has been involved in extensive research on the chemical and nutritive evaluation of corn and wheat DDGS and development of nutrient availability data for poultry and swine. To our knowledge, what has not yet been considered in the DDGS research is the fact that as co-products of brewer’s yeast (Saccharomyces cerevisiae) fermentation, they contain a significant quantity of yeast biomass which could be beneficial for gut development and health and effective in immune system stimulation. Based on our analysis, the residual yeast biomass content averaged 6.2 and 5.6% for wheat and corn DDGS, respectively. This could be of importance in light of a variety of Saccharomyces cerevisiae yeast-based products currently being offered to the poultry industry as growth promoters and natural alternatives to antibiotics (Stanley et al., 2004; Solis de los Santos et al., 2007). 
Table 5. Degradation of yeast cell wall polysaccharides following incubation of yeast-derived products with the yeast cell wall-lytic enzyme.
Enzyme Hydrolysis Products and Yeast-derived Products as Prebiotics and Natural Alternatives to Antibiotic Growth Promoters - Image 6
Interesting research data have recently been generated in our laboratory as a result of these studies. It appears evident that the use of a yeast cell wall lytic enzyme beta-1,3-glucan laminaripentaohydrolase from Arthobacter luteus can significantly depolymerize yeast cell wall polysaccharides so they become water-soluble and thus more bioactive. In addition, yeast cell lysis would result in the release of a variety of nutrients, including nucleotides, which may play a role in immune system development. It would appear that following enzyme treatment the mannan polysaccharides remain intact, but are released from the cell wall structure and thus become water-soluble. As illustrated in Table 5, the difference in yeast cell wall polysaccharide contents between the control and enzyme-treated samples indicates how much of the cell wall components and their hydrolysis products are present in the water phase and thus water-soluble. The yeast beta-1,3-glucans would now represent a variety of simple glucose, glucooligosaccharides and low-molecular weight polysaccharides of glucose while mannans would stay relatively intact, yet dissociated from the beta-glucan structure. 
Although this research is still in its infancy, a combination of an existing multicarbohydrase preparation fortified with a yeast cell wall lytic activity could serve as an effective and inexpensive alternative technology to replace antibiotic growth promoters in poultry and swine nutrition. 
Bedford, M. and J. Apajalahti. 2001. Microbial interactions in the response to exogenous enzyme utilization. Pages 299-314 in M. R. Bedford and G. G. Partridge, eds. Enzymes in farm animal nutrition CABI Publishing, Wallingford, UK.
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Fukata, T., Y. Hadate, E. Baba, and A. Arakawa. 1991. Influence of bacteria on Clostridium perfringens infections in young chickens. Avian Dis. 35:224-247.
Gibson, G. R. and M. B. Roberfroid. 1995. Dietary modulation of the human colonic microbiota: introducing the concept of prebiotics. J. Nutr. 125:1401-1412.
Huff, G. R., W. E. Huff, N. C. Rath, and G. Tellez. 2006. Limited treatment with beta-1,3/1,6-glucan improves production values of broiler chickens challenged with Escherichia coli. Poult. Sci. 85: 613-618.
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Jia, W., B. A. Slominski, H. L. Bruce, C. M. Nyachoti, and R. O. Jones. 2009. Enzyme addition facilitates the post-disease compensatory growth of broiler chickens challenged with Clostridium perfringens. Can. J. Anim. Sci. 89:369-381.
Jia, W., B. A. Slominski, H. L. Bruce, G. Blank, and O. Jones. 2009. Effects of diet type and enzyme addition on growth performance and gut health of broiler chickens during sub-clinical Clostridium perfringens challenge. Poult. Sci. 88:132-140.
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Kiarie, E.G., B.A. Slominski, D.O. Krause, and C.M. Nyachoti. 2008. Non-starch polysaccharides hydrolysis products of soybean and canola meal protect against enterotoxigenic Escherichia coli in piglets. J. Nutr. 138:502-508.
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Slominski, B.A. 2011. Recent advances in research on enzymes for poultry diets. Poult. Sci. 90:2013-2023.
Solis de los Santos, F. M. B. Farnell, G. Teliez, J. M. Balog, N. B. Anthony, A. Torres-Rodriquez, S. Higgins, B. M. Hargis, and A. M. Donoghue. 2005. Effet of prebiotic on gut development and ascites incidence of broilers reared in a hypoxic environment. Poult. Sci. 85:1092-1100.
Solis de los Santos, F., A. M. Donoghue, M. B. Farnell, G. R. Huff, W. E. Huff, and D. J. Donoghue. 2007. Gastrointestinal maturation in accelerated in turkey poults supplemented with a mannan-oligosaccharide yeast extract (Alphamune). Poult. Sci. 86:921-930.
Stanley, W.. G., C. Gray, M. Daley, W. F. Krueger, and A. E. Sefton. 2004. An alternative to antibiotic-based drugs in feed for enhancing performance of broilers grown on Eimeria spp.-infected litter. Poult. Sci. 83:39-44
Zdunczyk Z., J. Juskiewicz, J. Jankowski, E. Biedrzycka, and A. Koncicki. 2005. Metabolic response of the gastrointestinal tract of turkeys to diets with different levels of mannan-oligosaccharide. Poult. Sci. 84:903-909.
Zhang, A. W., B. D. Lee, S. K. Lee, G. H. An, K. B. Song, and C. H. Lee. 2005. Effects of yeast (Saccharomyces cerevisiae) cell components on growth performance, meat quality, and ileal mucosa development in broiler chickens. Poult. Sci. 84:1015-1021.
This paper was presented at the AMENA Congress, Puerto Vallarte, Mexico in October 2013.
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Dr. Bogdan Slominski
University of Manitoba
Dr Ashish Dhakarey
18 de marzo de 2014

I am a veterinary microbiologist and have experience of field trials and closely associated probiotics manufacturing and use in Pigs as well as in other species.
It is very effective in natural ways. Infact probiotics are beyond then digestion. It has so many functions.
We are also manufecturer of more then 70 probiotics and other preprations.

Dr Ashish

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