The authors are with the department of animal sciences at the University of Illinois.
Industrial strains of Saccharomyces cerevisiae are heterogeneous and used in several industries including baking, brewing, distilling and wine production (Sumner and Avery, 2002). Although these strains share common features such as efficient sugar utilization, high ethanol tolerance and production, high yield and fermentation rate and genetic stability, they also possess properties specific to each group (Trivedi et al., 1986; Benítez et al., 1996).
Eight official and one "tentative" yeast products are currently defined by the Association of American Feed Control Officials (AAFCO; 2002) and are differentiated by source of yeast and characteristics such as moisture and crude protein concentrations and fermentative activity. By definition, "brewers dried yeast" is the dried, non-fermentative, non-extracted yeast of the botanical classification Saccharomyces resulting as a byproduct from the brewing of beer and ale. It must contain not less than 35% crude protein and be labeled according to its crude protein content (AAFCO, 2002).
As defined, brewers dried yeast must originate from a brewery and the brewing of beverages -- beer or ale -- for human consumption and should not be confused with corn wet milling yeast that is used in industrial ethanol production.
As Table 1 illustrates, the chemical compositions of brewers dried yeast and corn wet milling yeast are different. Chemical and organoleptic differences likely are due to differences in the fermentation processes and in the substrates used. In the brewing industry, wort derived mostly from malted barley is fermented slowly at temperatures of 10-20 degrees C using a batch fermentation process that yields a beer with an alcohol content of approximately 6%.
In contrast, during wet milling ethanol fermentation, distillers commonly grind and cook corn using enzymes to convert starch to sugar. A rapid, continuous fermentation process at temperatures between 35 and 38 degrees C is then employed to maximize substrate utilization and ethanol production, yielding 9-12% alcohol.
Although not fully researched, many of the differences in brewers dried yeast and yeast from corn wet milling ethanol production (e.g., fat content, hops products [caryophyllene, humulene] and sugar profiles) are likely factors that affect palatability.
Composition of brewers yeast
Commercially available brewers yeast is typically dried from a yeast slurry to a dry powder of less than 10% moisture to facilitate handling, storage and transport. Brewers yeast is relatively high in crude protein and carbohydrate concentrations, while the concentrations of fat and ash are relatively low. This is not surprising because yeast synthesizes protein and vitamins while absorbing minerals from the beer wort during the fermentation process.
The relatively low fat content of brewers yeast compared to yeast from commercial wet milling ethanol fermentation likely is due to substrate differences (the low fat concentration of barley compared to corn) and differences in the fermentation processes.
Various methods are used to determine fiber concentration of ingredients, including brewers yeast. As Table 2 illustrates, the concentration of fiber present in yeast depends greatly on method used. Although the method of measuring crude fiber (AOAC, 1980) is used for regulatory purposes, results are misleading as several fibrous compounds are solubilized with this procedure, resulting in a large underestimation of fiber content.
The neutral detergent fiber (NDF) method of Robertson and Van Soest (1977) results in solubilization of viscous fiber components and recovery of cell wall constituents. Because brewers yeast contains a considerable amount of protein that becomes viscous when partially hydrolyzed during the NDF procedure, filtration problems and inflated recoveries result in overestimated fiber concentrations (Merchen et al., 1990). The best method for proteinaceous feeds such as brewers yeast is that of Prosky et al. (1992) used to measure total dietary fiber (TDF; Merchen et al., 1990).
The cell wall of S. cerevisiae constitutes approximately 15-30% of the dry weight of the cell and consists primarily of mannosylated proteins, beta-glucans and chitin (N-acetylglucosamine), which are covalently linked with one another. The glucan portion consists of beta-(1,3)- and beta-(1,6)-chains. The major structural components of the cell wall are the beta-(1,3)-glucans, which form the internal skeletal framework of the cell and are largely responsible for its mechanical strength. This form of glucan is highly branched and possesses multiple non-reducing ends that function as attachment sites for other components of the cell wall (Kollár et al., 1997). Beta-(1,6)-glucans are found primarily outside the skeletal framework and often are linked to cell wall proteins (CWP).
Mannose polysaccharides are linked to proteins to form a mannoprotein layer localized at the external surface of the yeast cell wall. Two classes of covalently linked CWP have been identified. The first class consists of glycosyl phosphatidylinositol (GPI) proteins, which form a GPI-CWP ® beta-(1,6)-glucan ® beta-(1,3)-glucan complex (Kollár et al., 1997).
The second class of CWP, the protein with internal repeats (Pir), are linked directly to beta-(1,3)-glucans. Mannoproteins are strictly regulated in response to changes in external conditions (e.g., heat shock, hypo-osmotic shock, carbon source) and internal changes during the cell division cycle (Horie and Isono, 2001).
While glucans and mannoproteins are main components of the cell wall and found in approximately equal amounts, chitin constitutes only about 1-3% of the cell wall. Although present in minute quantities, it is a major component of the primary septum and is involved in the separation of mother and daughter cells, making it essential for cell division (Shaw et al., 1991). The remaining components of yeast, excluding the cell wall, are collectively referred to as yeast cell extract and contain numerous nucleotides, enzymes, vitamins and minerals.
Use of yeast in pet foods
Brewers yeast is used in companion animal foods because it is a high quality protein source rich in B vitamins, amino acids and minerals. Inclusion of brewers yeast in companion animal diets has been shown to increase (P < 0.05) palatability in both dogs (Figure 1; Kennelwood Inc., unpublished data; Ontario Nutri Lab Inc., unpublished data) and cats (Figure 2; Kennelwood Inc., unpublished data) compared to diets containing wet milling yeast, with consumption ratios ranging from 1.9:1 to 2.1:1 in these experiments.
In these experiments, either brewers yeast or wet milling yeast was included in the diet at a 1% inclusion level. In each experiment, a panel of 20 dogs or cats was used to test food preference with a standard four-day palatability test. Each day on test, both diets were offered simultaneously for a period of one hour. To account for right-left bias, the placement of diets was alternated each day. After the one-hour feeding period, both diets were removed simultaneously and weighed to calculate intake. To our knowledge, more detailed nutritional studies with dogs and cats have not been conducted.
Use of yeast byproducts
In recent years, human food ingredients have been evaluated not only for their nutritional contribution but for their non-nutritional properties as well. "Functional foods," "nutraceuticals" and "phytochemicals" are terms commonly used to refer to foods or compounds in foods that possess properties that may benefit the human in ways other than providing nutritive value. Although use of functional foods began in the human food industry, the pet food industry, too, has recognized their potential benefits.
Many functional ingredients are thought to decrease the incidence of certain disease states or extend the lifespan of pets by possessing antioxidant activity, antimicrobial action or immuno-enhancing properties. Several components present in yeast may be classified as being "functional," including glucomannans, mannans, mannoproteins, beta-glucans and nucleotides.
Sharma and Márquez (2001) tested 12 pet foods commercially available in Mexico for frequency and concentration of aflatoxins. In that experiment, seven aflatoxins and aflatoxicol were detected in most samples, with aflatoxin B1 being present in the highest frequency and concentration. In all contaminated samples, maize was the main ingredient.
Research is needed to measure incidence and concentration of mycotoxins in pet foods commercially available in the U.S. and to determine whether these concentrations are cause for concern. If that is the case, inclusion of glucomannans in pet foods, especially those containing high concentrations of grain, may be prudent.
Due to the antimutagenic and antioxidative activity of glucomannans, they also could play a role in colon cancer prevention (Chorvatovicová et al., 1999; Križková et al., 2001).
Mannans have been studied for their ability to agglutinate and interfere with intestinal binding and colonization of harmful microbial species. Numerous Escherichia coli and Salmonella strains possess mannose-specific fimbriae, agglutinate mannans in vitro and colonize in lower concentrations in animals supplemented with mannans. Fimbrial adhesins specific for mannan residues are referred to as "Type-1" adhesins.
Mannans aid in the resistance of pathogenic colonization by acting as receptor analogues for Type-1 fimbriae and decrease the number of available binding sites (Oyofo et al., 1989).
Mannans are capable of modulating the immune system and influencing microbial populations in the gut. Mannans have been reported to increase (P = 0.14) serum immunoglobulin A (IgA) concentrations in dogs (2.33 versus 1.93 g/L; Swanson et al., 2002a). In adult dogs, MOS beneficially altered microbial ecology by increasing (P = 0.13) lactobacilli populations (9.16 versus 8.48 log10 colony-forming units [cfu]/g fecal dry matter) and decreasing (P = 0.05) total aerobe populations (7.68 versus 8.67 log10 cfu/g fecal dry matter; Swanson et al., 2002a).
An experiment using free-living, obese, hypercholesterolemic men demonstrated that yeast-derived beta-glucans were well tolerated and decreased (P < 0.05) blood total cholesterol concentrations similar to the effect of oat products (Nicolosi et al., 1999). In that experiment, beta-glucan supplementation decreased blood total cholesterol after seven (8% reduction) and eight (6% reduction) weeks of supplementation.
Yeast-derived beta-glucans also appear to possess antimicrobial and antitumor properties by enhancing immune function. The binding of beta-glucan to its receptor present on macrophages results in phagocytosis, respiratory bursts and secretion of tumor necrosis factor-alpha (TNF-alpha; Chen and Hasumi, 1993; Lee et al., 2001).
Finally, beta-glucans are readily fermented in the large bowel and serve as a fuel source for microbial populations.
In addition to stimulating the development of the small intestine (Bueno et al., 1994) and liver (Sánchez-Pozo et al., 1998), exogenous nucleotides have been shown to enhance immune function by increasing production of immunoglobulins, improving response to vaccines and increasing tolerance to dietary antigens (Maldonado et al., 2001).
Because of its importance in neonatal nutrition, the inclusion of nucleotides in human infant formulas is under investigation (Cordle et al., 2002; Ostrom et al., 2002).
In vitro research on yeast byproducts.
When using canine fecal inoculum to determine the fermentability characteristics of MOS, Vickers et al. (2001) observed the production of moderate concentrations of total short-chain fatty acids after 6 (0.49 mmol/g of organic matter), 12 (1.45 mmol) and 24 hours (2.40 mmol) of in vitro fermentation. Very low concentrations of lactate were produced as a result of MOS fermentation. The microbial species responsible for MOS breakdown were not determined in this experiment.
Hussein and Healy (2001) also performed an in vitro experiment using canine and feline fecal inoculum to determine fermentability of MOS. Differences were not observed in fermentability between dog and cat fecal inoculum. By examining dry matter and organic matter disappearance, it appeared that MOS was highly fermented. Dry matter disappearance after 6, 12, 18 and 24 hours of in vitro fermentation was 54.3, 57.9, 60.7 and 61.3%, respectively. Organic matter disappearance was similar to that of dry matter (56.8, 60.7, 63.7 and 64.1% after 6, 12, 18 and 24 hours of fermentation).
Dry matter and organic matter disappearance do not always reflect microbial fermentation due to the disappearance of soluble carbohydrates present in the substrates that are not retained during filtering. Although soluble carbohydrates are available for fermentation, gravimetric methods cannot determine the proportion used by the microbes as an energy source. Therefore, the measurement of dry matter and organic matter disappearance is not as accurate as the measurement of the fermentation end products (i.e., short-chain fatty acids and gas), which is a direct measurement of fermentation.
Concentrations of total short-chain fatty acids, acetate and propionate increased linearly over time. Moderate concentrations of total short-chain fatty acids (10.1, 26.8, 36.7 and 49.7 mM) were produced after 6, 12, 18 and 24 hours. In comparison to total short-chain fatty acids, lactate concentrations were fairly high (7.7, 8.7, 7.6 and 5.9 mM), suggesting fermentation by a lactate-producing species (e.g., lactobacilli, bifidobacteria).
In agreement with the work of Vickers et al. (2001), these data suggest that MOS is moderately fermentable by canine and feline microflora. The lactate produced during fermentation suggests that lactate-producing species are able to utilize MOS, possibly by acting as a prebiotic for these species.
In agreement with short-chain fatty acid data, gas was produced in relatively high amounts (corrected gas values were 17.4, 58.3 and 100.9 ml/g organic matter after 4, 12 and 24 hours of fermentation).
Canine research on yeast byproducts.
In the second experiment, Border collie pups were fed diets containing 0 or 2 g MOS/kg. After a seven-day adaptation period, a vaccination protocol was initiated. All dogs were vaccinated against parvovirus, leptospirosis, adenovirus and distemper. Vaccine boosters were applied on day 21 for leptospirosis and on day 35 for parvovirus. Blood characteristics were measured over a nine-week period.
No changes were observed in weight gain, lysozyme activity, plasma protein concentration or plasma IgG concentration. Neutrophil activity was numerically increased in pups fed the diet containing MOS after vaccination (approximately 18 versus 14 Nitroblue tetrazolium [NBT]+ cells per slide]. However, due to low animal numbers (n = three per group), statistical significance was not reached.
Using adult ileal cannulated dogs, Strickling et al. (2000) compared a control diet to those containing 5 g oligosaccharide/kg diet, one of which was MOS. Researchers measured ileal and total tract nutrient digestibilities, microbial populations, ileal pH, ammonia and short-chain fatty acid concentrations, blood glucose and fecal consistency. Besides minor changes in short-chain fatty acid concentrations, the only relevant finding was a decrease (P = 0.07) in Clostridium perfringens populations in dogs fed MOS (4.48 log10 cfu/g) versus dogs fed xylooligosaccharides (5.16 log10 cfu/g) or oligofructose (4.74 log10 cfu/g).
Because clostridia species do not possess mannose-specific fimbriae, another mechanism is likely occurring. The lack of any significant findings may be due to the low dose of prebiotics consumed (only about 1.3 g per day) or to the use of soybean meal in the control diet, which supplied an estimated 10 g/kg of naturally occurring oligosaccharides, mainly galactooligosaccharides. Any beneficial effects resulting from MOS consumption may have been masked by the presence of these naturally occurring oligosaccharides.
Zentek et al. (2002) used four dogs in a 4 x 4 Latin square design to determine the effects of MOS, transgalactosylated oligosaccharides, lactose and lactulose on fecal characteristics, total tract digestibility and concentrations of microbial end products in feces and urine. Carbohydrate supplements were administered at a rate of 1 g/kg bodyweight per day. MOS supplementation decreased (P < 0.05) fecal pH (6.6 versus 6.9), fecal ammonia excretion (78.4 versus 116 ;Jmol/g feces) and apparent dry matter (81.9 versus 85.0%), crude protein (79.8 versus 82.5%) and nitrogen-free extract (83.1 versus 94.8%) digestibilities.
By decreasing fecal pH and ammonia, MOS supplementation appeared to improve indices of colonic health. However, the decreases observed in apparent nutrient digestibilities resulting from MOS supplementation would increase fecal quantity and the cost of feeding the animal. The dose of carbohydrate supplements fed in this experiment (1 g/kg bodyweight per day) was very high. Smaller doses of MOS may not have such negative effects on nutrient digestibility.
Using ileal cannulated adult dogs, Swanson et al. (2002a) examined the effects of supplemental MOS and/or fructooligosaccharides (FOS) on colonic microbial populations, local and systemic immune function, fecal protein catabolite concentrations and ileal and total tract nutrient digestibilities. A 4 x 4 Latin square design with 14-day periods was used. Twice daily, dogs were offered 200 g of a dry, extruded kibble diet and given the following treatments orally via gelatin capsules: (1) control (no supplemental MOS or FOS), (2) 1 g FOS, (3) 1 g MOS or (4) 1 g FOS and 1 g MOS.
MOS supplementation beneficially influenced microbial populations, decreasing (P = 0.05) total aerobe (7.68 versus 8.67 log10 cfu/g fecal dry matter) and tending to increase (P = 0.13) lactobacillus populations (9.16 versus 8.48 log10 cfu/g fecal dry matter). MOS also increased serum IgA concentrations (2.33 versus 1.93 g/l, P = 0.14) and lymphocyte numbers (20.4 versus 15.6% of total white blood cells, P < 0.05). A tendency for decreased ileal dry matter (55.0 versus 67.7 %, P = 0.15) and organic matter (63.6 versus 74.1%, P = 0.15) digestibility also was observed from MOS supplementation.
The combination of FOS and MOS supplementation enhanced immune characteristics, increasing ileal IgA concentrations on a dry matter basis (4.90 versus 3.40 mg/g ileal dry matter, P = 0.06) and crude protein basis (12.22 versus 8.22 mg/g ileal crude protein, P = 0.05). Supplementation of FOS plus MOS also decreased (P < 0.05) total fecal indole and phenol concentrations (1.54 versus 3.03 ;Jmol/g fecal dry matter), compounds partially responsible for fecal odor and detrimental to intestinal health.
This experiment was performed using healthy adult dogs, which would not be at the highest risk for intestinal irregularities. It is likely that the health benefits of feeding MOS alone, or in combination with FOS, would be more beneficial to populations of elderly dogs, young weanling puppies or stressed animals.
In a follow-up study, Swanson et al. (2002b) supplemented ileal cannulated dogs with either 1 g sucrose (placebo) or 2 g FOS plus 1 g MOS. Fecal, ileal and blood samples were collected at the end of each 14-day period to measure microbial populations and immune characteristics. Supplementation of FOS plus MOS increased (P < 0.05) fecal bifidobacteria (10.04 versus 9.42 log10 cfu/g fecal dry matter) and lactobacilli concentrations in feces (9.75 versus 8.24 log10 cfu/g fecal dry matter) and ileal effluent (8.66 versus 7.55 log10 cfu/g ileal dry matter).
Dogs fed FOS plus MOS also tended to have lower (P = 0.08) blood neutrophils (62.99 versus 66.13% of total white blood cells; 6.40 versus 7.22 x 103 cells/;Jl) and greater (P = 0.06) blood lymphocytes (19.95 versus 17.29% of total white blood cells) compared to placebo. Serum, fecal and ileal immunoglobulin concentrations were unchanged (P >0.05) by treatment. Supplementation of FOS plus MOS beneficially influenced indices of gut health by improving ileal and fecal microbial ecology and altered immune function by causing a shift in blood immune cells.
Active components in yeast byproducts?
Source B was unique in that it contained a considerable amount of galactose in addition to glucose and mannose. The presence of galactose in that source may suggest that guar gum or locust bean gum, which contain galactomannans, are also present in this source of MOS. Although marketed as a source of MOS, these products are very complex and also contain glucans, mannoproteins, phosphate and several other compounds that apparently are not excluded in the crude extraction process.
Because the composition of MOS is complex, the components that result in beneficial effects are not known. Although the mannan portion of MOS is generally thought to be responsible for the pathogenic resistance effect by acting as a receptor analog for Type-1 fimbrial adhesions present on species such as E. coli and salmonella, it is possible that a different fraction present in MOS is responsible for its effects on immune function.
For example, mannoproteins and beta-glucans taken from yeast cell walls have been reported to enhance immunity. Therefore, more research is needed in order to determine whether bioactive peptides, beta-glucans, mannans or unknown factors present in MOS are responsible for the immune responses observed as a result of their supplementation.
Relatively little research has been performed with companion animal species, but the limited data suggest that inclusion of brewers yeast or yeast byproducts in pet foods may support gut health. Although brewers yeast often is included for palatability enhancement, its functional properties may be an even more important reason for its inclusion in pet foods.
From the limited number of experiments testing MOS, it appears that it has beneficial effects on indices associated with gut health. MOS supplementation has resulted in improved ileal and fecal microbial ecology and enhanced immune status. Because glucomannans are able to bind mycotoxins, their presence in pet foods also may be beneficial. Finally, yeast-derived beta-glucans, mannoproteins and nucleotides require further testing to determine their role, if any, in companion animal nutrition and health.
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