Cellulase Supplementation Does Not Improve the Digestibility of a High-Forage Diet in Horses

REFEREED

ORIGINAL RESEARCH

Cellulase Supplementation Does Not Improve the Digestibility of a High-Forage Diet in Horses C.I. O’Connor-Robison, MS, B.D. Nielsen, MS, PhD, and R. Morris, BS ABSTRACT Six mature Arabian geldings were used in a two-period crossover study to investigate the effects of cellulase supplementation on fiber digestion. Horses were randomly assigned to either a control (CO; n ¼ 3) or a cellulase (CE; n ¼ 3) treatment for the first period and then treatments were switched for period 2. Each period consisted of a 10-day diet adaptation followed by a 3-day total fecal collection. The enzyme mixture contained 40,000 cellulase units/g and was fed at a rate of 3 g/day split evenly between two feedings. During the diet adaptation period, horses had ad libitum access to timothy hay and were also fed 165 g whole oats as a carrier for the supplement. When eating the CO treatment, horses consumed 16% more hay than when on the CE treatment (P ¼ .004). Fecal output also tended to be greater when horses consumed the CO treatment as compared with CE treatment (P ¼ .07). No differences were found between treatments for fecal percent dry matter (DM%), fecal neutral detergent fiber (NDF), fecal acid detergent fiber (ADF), fecal nitrogen (N), or fecal gross energy (GE). There was a trend for horses consuming the CO treatment to digest more NDF than when consuming the CE treatment (34.6%  1.5 vs 31%  1.5; P ¼ .07). Horses also digested a greater %ADF, %N, and Mcal of energy when consuming the CO treatment than when consuming the CE treatment (P < .05). Cellulase addition to a hay-based horse diet decreased digestion of fiber components. Keywords: Cellulase; enzyme

Fiber

digestion;

Fibrolytic

INTRODUCTION Crude fiber is composed of three major constituents: cellulose, hemicellulose, and lignin.1 Crude fiber has been thought of as the indigestible portion of animal feed because mammals do not produce enzymes capable of breaking down crude fiber. However, mammals are able to use the substrates produced by the breakdown of crude fiber by microbial fermentation for energy. These microflora From the Department of Animal Science, Michigan State University, East Lansing, MI. Reprint requests: Cara O’Connor-Robison, MS, 1287 Anthony Hall, East Lansing, MI 48824. 0737-0806/$ - see front matter Ó 2007 Elsevier Inc. All rights reserved. doi:10.1016/j.jevs.2007.10.014

Journal of Equine Veterinary Science  Vol 27, No 12 (2007)

are found in the rumen of ruminant animals and in the cecum and colon of nonruminant herbivores such as horses and rabbits. These microorganisms produce the enzymes, such as cellulase, necessary to break down fiber. Supplementation of cellulase and other fibrolytic enzymes into animal diets has increased in popularity in recent years as a result of environmental pressures to reduce nutrient excretion.2 Cellulase is marketed across the United States, often as part of multi-enzyme complexes. In addition to cellulase, these multi-enzyme complexes may contain digestive enzymes such as amylase, protease, lipase, and xylanase. Numerous studies to investigate the potential benefit of exogenous digestive enzyme supplementation have been completed in ruminants. Supplementation of xylanases and cellulases to ruminant diets have been shown to improve total tract, neutral detergent fiber (NDF), and fecal acid detergent fiber (ADF) digestibility.2 Bowman et al3 observed that when xylanases and cellulases were applied to the concentrate portion of the ration, dairy cows had a higher dry matter (DM) and organic matter total tract digestion. According to Beauchemin and coworkers,4 the addition of fibrolytic enzymes to both high-grain and high-forage diets fed to ruminants can improve fiber digestibility. Although a substantial amount of research has been done to investigate the effects of exogenous enzyme administration to ruminant diets, little research has been done to investigate the same in monogastric hindgut-fermenters such as rabbits and horses. Eiben et al5 fed a cellulasehemicellulase enzyme complex to rabbits, resulting in improved feed efficiency and decreased mortality because of enteritis. In horses fed a diet of grass hay supplemented with either textured concentrate, pelleted concentrate, whole oats, or alfalfa cubes, cellulase administration improved the digestion of NDF and ADF in the oat and textured feedsupplemented diets, but decreased digestion of NDF and ADF in the alfalfa-supplemented diet.6 Clearly, further research is needed to identify benefits from feeding supplementary fibrolytic enzymes, such as cellulase, to the horse. The hypothesis of this study was that feeding horses a cellulase supplement in addition to their regular diet would improve fiber digestion.

MATERIALS AND METHODS Six mature Arabian geldings (12  0.6 years of age) weighing 449  13 kg were used in a two-period crossover 535

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536

Table 1. Body Weights during the Adaptation Periods and Total Collections Treatment

Adaptation 1

Fecal Collection 1

Adaptation 2

Fecal Collection 2

End

Cellulase supplement (CE) Control supplement (CO)

433  18

465  18

474  18

474  18

467  18

465  18

478  18

458  18

464  18

460  18

design study. Horses were housed in individual 2.7  3.4-m box stalls bedded with shavings and exercised at 3 m/second for 30 minutes 5 days/week in a free flow exerciser (Equicizer, Santa Monica, CA). The experimental design and all procedures were approved by the All University Committee on Animal Use and Care of Michigan State University. Before the start of the study, all horses were housed on pasture together and received no supplemental feed. Before the start of the first period, horses were randomly assigned to either a control (CO; n ¼ 3) or a cellulase (CE; n ¼ 3) group. The CO supplement was a placebo formulated to simulate the treatment and contained 42% red wheat bran, 20% wheat germ, 19% baker’s flour, and 19% evaporated cane juice crystals (Winner’s Bee Pollen). The CE supplement (Cellulase Supplement, Winner’s Bee Pollen) contained the same ingredients with the addition of 40,000 cellulase units/g and was fed at a rate of 3 g/ d split evenly between two feedings. Both the CO and CE supplements were loose and not pelleted. Researchers were blinded to treatments until data analysis was completed. Horses were fed 165 g whole oats mixed with 1.5 g either the CE or CO supplement in a bucket at 7:30 AM and 4:00 PM. Additionally, horses had ad libitum access to second-cutting timothy grass hay. Hay was provided in wall-mounted hay feeders during adaptation periods and in hay nets during the total fecal collection. Horses had free access to fresh water and trace mineral blocks at all times. During period 1, horses underwent a 10-day diet adaptation followed by a 3-day total fecal collection. During period 2, horses switched treatments and underwent another 10-day diet adaptation followed by another 3-day total fecal collection. During the fecal collection, hay intake was measured at 7:30 AM and 4:00 PM, and horses were fed 5% over the amount consumed during the previous day for the same period. During each period, samples of hay and oats were collected at each feeding time via grab samples throughout the fecal collection. Body weight was measured at the start of the study, and at the beginning of each 3-day total collection period on an electronic scale (Table 1). During the total fecal collection, feces were collected from the total collection devices (Equisan, Australia) every 8 hours. At each collection time, total feces were weighed, and a 10% sample was saved and frozen for later analyses. After completion of the study, all of the feces from each horse/period combination were thawed and pooled, and 0.5-kg samples were retained and frozen at 208C until

they could be freeze-dried. After samples were freezedried, the fecal matter along with the hay, oats, and both supplements were ground through a cyclone mill with a 0.5-mm screen. Hay, oats, and supplements were ground and analyzed on an as-fed basis, but are reported on a DM basis. Feces, hay, oats, and supplements were analyzed for percent DM, NDF, ADF, N, and gross energy (GE). Oven dry matters were determined for feces before and after freezedrying and for hay, oats, and supplements in triplicate. All analyses were conducted using the freeze-dried fecal samples. The analyses of ADF and NDF were completed using the Van Soest method for fiber analysis.7 Nitrogen content was analyzed with a Leco FP–2000 (St. Joseph, MI) using AOAC Method 990.03. GE contents of feeds and feces were analyzed using a bomb calorimeter (Parr, model 1242, Moline, IL). The DM, NDF, ADF, crude protein (CP), and GE values for oats, timothy hay, and the two supplements (CE and CO) are presented in percentages and mg/kg body weight (BW) in Table 2 on a dry matter basis. Statistical Analysis Data were analyzed using the MIXED procedure of SAS 8.2.8 Period and horse were considered random effects, and treatment was considered a fixed effect. Differences were considered significant at P < .05, and trends were discussed at P < .1. All results are reported as means  standard error of the mean (SEM).

RESULTS There was no difference between the body weights of horses while on the CE and CO treatments at any time (treatment effect, P ¼ .63; Table 1). On a g/kg BW DM basis, the CO treatment consumed 16% more hay (15.9  1.4 g/kg BW) than CE (13.3  1.4 g/kg; P ¼ .004). The CO treatment tended to have a higher average daily fecal output compared with the CE treatment. There were no differences between treatments for fecal DM percentage, percentage fecal NDF, percentage fecal ADF, percentage fecal N, or fecal GE (Table 3). Feed Digestibility The CO treatment tended to have a lower total tract digestibility than the CE treatment (P < .10; Table 3). There was a trend for horses on the CO treatment to digest a greater percentage of NDF than when on the CE

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Table 2. Percentages of DM, NDF, ADF, CP, and GE Values of Feeds and Supplements Used throughout the Trial on a Dry Matter Basis Item

Oats

Timothy Hay

Cellulase Supplement (CE)

Control Supplement (CO)

% DM % NDF % ADF % CP GE (kcal/g) Amt. consumed(g/kg BW) DM (g/kg BW) NDF (g/kg BW) ADF (g/kg BW) N (g/kg BW) GE (kcal/kg BW)

88.6% 29.7% 13.0% 12.5% 4,552 0.70 0.62 0.18 0.081 0.014 2.8

88.5% 68.3% 43.8% 8.9% 4,420 16.6 14.7 10.05 6.44 0.236 65.0

92.1% 18.5% 6.6% 16.0 % 4,349 0.0064 0.0059 0.0011 0.0004 0.16 0.026

92.1% 21.0% 7.2% 16.6% 4,244 0.0064 0.0059 0.0011 0.0004 0.17 0.025

treatment (P < .10). The CO treatment also digested more ADF and more energy than the CE treatment (P < .05). Additionally, the CO-treated horses tended to digest more N when compared with those treated with CE (P < .10).

DISCUSSION The results from this study indicate that addition of cellulase to the concentrate portion of a horse’s diet decreased dry matter intake and fiber digestibility. Bowman et al3 reported that, in cattle, the portion of the diet to which the enzyme supplement is added plays an important role in determining whether digestibility will be affected. When enzymes were added to the larger portion of the diet, they were more effective in increasing fiber digestion than when added to a small portion of the diet.3 The authors speculated that adding the enzyme to a large portion of the diet maximized dispersion in the rumen, whereas adding the enzyme to a small portion increased the rate of passage of the enzyme through the digestive tract.3 In the current study, the enzyme was added to 165 g whole oats, which were used as a carrier, and this small amount would not likely impact rate of passage. A study by Beauchemin et al.4 reported that certain enzymes are more effective than others for degradation of a certain feed. In the current study, cellulase may not have been the best enzyme for increasing digestibility of a timothy haybased diet. Murray et al9 examined an enzyme product containing 6,222 IU cellulase, 1,039 IU xylanase, and 2,156 IU b-glucanase and found that when applied at 2.4 ml/g to lucerne hay and degraded in vitro with a simulated foregut digestion, the pre-cecal degradability of nonstarch polysaccharides increased. Perhaps more than one enzyme needs to be present to elicit a positive change in feed digestibility in higher-fiber diets. When supplementing xylanases to beef cattle, more consistent results have been shown when the enzyme has been added to high-concentrate versus high-forage diets.4 This is

Table 3. Fecal Output and Total Tract Digestibility for Horses Fed either a Control Diet (CO) or a Diet Containing Cellulase (CE) on a DM Basis Item

CO

CE

SEM

P Value

Fecal output (kg DM) Fecal DM (%) Fecal NDF (%) Fecal ADF (%) Fecal N (%) Fecal GE (Mcal) % DM digested % NDF digested % ADF digested % N digested % Energy digested

4.5

4.2

0.1

.07

20.9 72.1 47.7 7.2 4,330 57.9 34.6 32.1 71.0 40.7

20.6 71.8 47.8 7.3 4,275 55.7 31.0 27.8 69.1 34.3

0.3 0.3 0.3 0.2 36.3 1.4 1.5 1.5 0.8 1.3

.33 .35 .71 .48 .20 0.06 0.07 0.04 0.07 0.008

also true when horses are supplemented with fibrolytic enzymes on either high-grain or high-forage diets, because enzyme addition increased DM and fiber digestibility only in the high-grain diets.6 Nsereko et al10 demonstrated that there is a negative correlation between concentrations of exogenous b-linked glucan hydrolases, such as cellulase, and fiber digestion in the rumen, suggesting that the hydrolases may block enzyme sites needed by microbial enzymes. In a study by Murray et al,11 ponies fed low concentrations of enzyme-treated forages had decreased apparent digestibility of organic matter and NDF. It was hypothesized that the enzymes were at too low of a concentration to break down the fiber in the hay but were able to block microbial binding sites.11 If this is true, it may help explain why overall fiber digestion was decreased in the CE horses. Nsereko et al10 concluded that feeding a solid enzyme preparation without pre-incubation with the feedstuff would not have a beneficial effect on fiber digestion and recognized that a pre-ingestion feedenzyme interaction must take place. Supplementation of fibrolytic enzymes to

538

horses may be more effective if the feedstuffs were pre-incubated with enzyme before ingestion, although doing so is not practical under most feeding management programs. In conclusion, cellulase addition to a hay-based horse diet did not increase the digestion of NDF and appeared to actually decrease digestibility of several dietary components. Although a number of theories exist to explain this, it may be warranted to investigate the mode of enzyme action further in the cecum and colon of horses. Additionally, although this is only one study that has found that adding cellulase to a small component of a horse’s diet has a negative effect on nutrient utilization, it does suggest that supplementation of cellulase in commercial concentrate preparations should be avoided until further research is conducted. REFERENCES

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