Estimation of Total Tract Apparent Digestibility of Nutrients in Three Annual Winter Forages Using Two Different Digestibility Markers D. Henry, C. Njombwa, F. Ciriaco, V. Mercadante, G. Lamb, and N. DiLorenzo 1 When determining total tract digestibility of annual winter forages such as ryegrass, oat, and triticale, utilization of either titanium dioxide or chromic oxide were equally accurate estimators of digestibility. In addition, collection of fecal samples two or three per day also did not differ in their ability to predict total tract digestibility. Summary A study was performed to: 1) compare total tract digestibility of nutrients for three annual winter forages (ryegrass (Lolium multiflorum Lam.)), ryegrass + oat (Avena sativa), and ryegrass + triticale (Triticosecale rimpau) and 2) determining the efficacy of two digestibility markers: titanium dioxide (TiO 2 ) and chromic oxide (Cr 2 O 3 ) used with a fecal sample collection protocol of either 2 or 3 /d. Twelve Angus and Angus crossbred heifers were utilized in pens of 2 within the Feed Efficiency Facility in Marianna, Fl. Heifers were fed daily fresh cuttings of 1 of the 3 treatment forages ad libitum. A GrowSafe System (GrowSafe Systems Ltd., Alberta, Canada) was used to determine feed intake which was used to evaluate dry matter (DM), organic matter (OM), crude protein (CP), neutral detergent fiber (NDF), and acid detergent fiber (ADF) intake of heifers. There was no difference (P > 0.05) between the 3 forages for DM, OM, CP, NDF, and ADF intake. No difference (P > 0.05) was noted for total tract digestibility of the three forages. When evaluating markers and fecal collections, no differences (P > 0.05) were found between the different fecal collection protocols or the 2 digestibility markers. When comparing markers, both TiO 2 and Cr 2 O 3 can be used interchangeably along with fecal collections at 0800 and 1600 h. Introduction The digestibility of a feedstuff is a very good indicator of its nutritive value. Measuring digestibility allows researchers to determine the extent to which a nutrient is absorbed within an animal and thus, decide how much of these nutrients are available for maintenance, growth, reproduction, and production of protein such as milk and meat. As it would imply, feeds with high digestibility have a tendency to have higher nutritive values, whereas feeds with low digestibility have a lower nutritive value (Kham et al., 2003). A livestock producer can potentially increase the efficiency of their operation by choosing forage varieties, grains or byproducts, based on the digestibility of the feedstuffs available for consumption by the animals. There are multiple techniques available to evaluate digestibility of feeds. The use of substances that are indigestible within the digestive system of an animal (called digestibility markers) is only one of the methods. Characteristics of a good digestibility marker are: 1) they are strictly non-absorbable; 2) they do not affect nor are they affected by the digestive system or microbial population; 3) they are physically similar to or closely associated with feed material; and 4) methods of estimation in digesta samples must be specific, sensitive, and not interfere with other analyses (Kham et al., 2003). Sunvold and Cochran (1991) found that some markers produced a very accurate estimate of digestibility for some feeds while giving less than desirable results for other feeds. With this inconsistency of feed evaluation, it can be difficult to determine the correct nutritive value of a feed or forage needed to meet maintenance and production requirements of livestock. In the past, Cr 2 O 3 has been intensively used as a digestibility marker. Nevertheless, there have 1 North Florida Research and Education Center, University of Florida, Marianna, FL
been reports that describe carcinogenic effects of Cr 2 O 3 (Myers et al., 2004). As a alternative, TiO 2 is currently being explored as a comparable marker. Previous studies state the effectiveness of TiO 2 as a marker in rats, chickens, pigs, and dairy and beef cattle (Myers et al., 2004 and Titgemeyer et al., 2001). Currently, there is no available data to assess the viability of using titanium dioxide as a digestibility marker in beef cattle fed fresh forages on an ad libitum basis, or to quantify the effect of collecting fecal samples 2 or 3 /d. Therefore, the efficacy of TiO 2 as a marker for these conditions is unknown, limiting its application as a digestibility marker in experiments. Therefore, a study was conducted with objectives to: 1) compare total tract digestibility of nutrients for three cool-season forages, and 2) to determine the efficacy of two digestibility markers, TiO 2 and Cr 2 O 3 sampled 2 or 3 /d on estimating digestibility. Ryegrass (Lolium multiflorum Lam.), ryegrass combined with oat (Avena sativa), and ryegrass combined with triticale (Triticosecale rimpau) were the coolseason forages evaluated. Materials and Methods Ryegrass (Lolium multiflorum Lam cultivar Prine; R), a blend of ryegrass (Lolium multiflorum Lam cultivar Prine) and oat (Lolium multiflorum Lam cultivar Prine; OR), and a blend of Ryegrass (Lolium multiflorum Lam cultivar Prine) and triticale (Triticosecale rimpau cultivar Trical 342; TR) were the forages utilized in this study. These forages were sown into prepared seedbeds in pastures of 1.75 ac. The forages were sown at a rate of: 31 lbs/ac of ryegrass; 60 lbs/ac of oat combined with ryegrass sown at the rate of 27 lbs/ac; and triticale was sown at the rate of 244 lbs/ac in combination with ryegrass at 15 lbs/ac. All pastures were fertilized twice; the first fertilization was done 28 d after planting at the rate of 50 lbs/ac of N (NH 4 NO 3 ) and 19 lb/ac of S while the second was done after another 28 d from the first fertilization at the rate of 50 lb/ac of N (NH 4 NO 3 ) and 10 lb/ac of S. Forages were allowed 98 d from planting prior to harvesting. Herbage mass estimates were made at the beginning of the study by forage clippings of a 2.69 feet 2 surface in the pasture and are reported in Table 1. In addition, DM, CP, NDF, and ADF concentration of the forages are reported in Table 1. Crude protein was analyzed by a macro nitrogen analyzer (Vario Max CN, Elementar Americas Inc., Mt. Laurel, MJ) and NDF and ADF analysis was performed using an Ankom 200 Fiber Analyzer (Ankom Technology, Fairport, NY). This study was performed using 12 Angus and Angus crossbred heifers. The heifers were randomly assigned to 1 of the 3 forage treatments (R, OR, or TR). After grazing for 28 d, the heifers were weighed on d 29 (801 ± 114 lbs of body weight [BW]), arranged by BW, and assigned to pens (2 heifers/pen), randomly, in the Feed Efficiency Facility (FEF) at the North Florida Research and Education Center (NFREC) in Marianna, FL. Ad libitum access to water and daily, fresh cut forage of the same treatments the heifers were already consuming were given from d 29 to d 44. Individual forage intake was measured in the FEF using a GrowSafe System (GrowSafe Systems Ltd., Alberta, Canada). While in the FEF, the heifers were given 2 digestibility markers via gelatin capsules daily. Daily, 10 g of TiO 2 and 10 g of Cr 2 O 3 were administered orally by a balling gun. Titanium dioxide was analyzed as described by Myers et al. (2004). The method of Williams et al. (1962) was utilized to digest and analyze Cr 2 O 3. Feed samples were collected daily from d 39 to d 43 and fecal samples, collected by rectum grabs, were taken from d 40 to d 44. Daily DM intake measurements were determined by taking the average of 2 daily feed samples at 8:00 a.m. and 4:00 p.m. Feed samples were dried at 212 F (100 C) for 24 h to obtain a DM weight. Feed samples used for nutritive value (CP, ADF, and NDF) were collected once daily and frozen at -4 F (4 C) until being freeze dried. After the freeze drying process, samples were ground through a 2- mmscreen before analysis. Three samples of feces were taken each d for 5 d at 8:00 a.m., 12:00 p.m., and 4:00 p.m. To determine if there is a difference between collecting feces 2x and
3x/d, composites of each heifer were made for 8:00 a.m. and 4:00 p.m., as well as 8:00 a.m., 12:00 p.m., and 4:00 p.m. For this study a split-split plot design in which the whole plot tested the forage treatment effect, the split plot tested the fecal collection schedule (2x vs. 3x) and the split-split plot tested the marker effect (Cr 2 O 3 vs. TiO 2 ) using heifer as the experimental unit. The data was analyzed using the MIXED procedure of SAS. All values reported are least square means and significance was declared if P < 0.05. Results Daily nutrient intake of the heifers was estimated during the study (Table 2). There was no difference (P > 0.05) between the 3 forages for DM, OM, CP, NDF, and ADF intake. The intakes of DM, OM, CP, NDF, and ADF were 10 lbs/d, 8.9 lbs/d, 1.7 lbs/d, 4.5 lbs/d and 2.2 lbs/d, respectively. No difference (P > 0.05) was noted for total tract digestibility of the 3 forages. On average for the 3 forages (R, OR, and TR), the DM, OM, CP, NDF, and ADF digestibility in the total tract were 59%, 61%, 60%, 49%, and 35%, respectively. Neither the different fecal collection protocols nor the 2 digestibility markers had significant effects (P > 0.05) on total tract digestibility (Table 3). There were no significant differences (P > 0.05) within the TiO 2 analysis for the 2 fecal collection protocols (2 and 3 daily). Similarly, no difference (P > 0.05) was found within the Cr 2 O 3 for the 2 fecal collection procedures. No interactions (P > 0.05) were found between the markers and forage. Conclusion As indigestible markers, both TiO 2 and Cr 2 O 3 can be used to evaluate the digestibility of winter forages. When measuring the total tract digestibility of winter forages, it is sufficient to collect fecal samples only twice per d (8:00 a.m. and 4:00 p.m.). This eliminates further stress placed upon the cattle by removing the 12:00 p.m. handling to collect another fecal sample. Overall, when comparing markers, both TiO 2 and Cr 2 O 3 can be used interchangeably along with fecal collections in the morning and afternoon. Further research is needed with digestibility markers such as internal markers to compare with the data presented in this report. Literature Cited Kham et al., 2003. Int. J. Agri. Biol. 5:1560. Myers et al., 2004. J. Anim. Sci. 82:179. Sunvold and Cochran, 1991. J. Anim. Sci. 69:4951 Titgemeyer et al., 2001. J. Anim. Sci. 79:10593. Williams et al., 1962. J. Agr. Sci. 59:381.
Table 1. Composition, yield and analyzed nutrient content of the winter annual pastures used. Treatment 1 Item Ryegrass 2 ryegrass 3 ryegrass 4 Oat + Triticale + Herbage mass at the beginning of digestibility phase, lbs DM/ac 2,012 3,086 4,631 Botanical composition, % of total herbage mass DM 5 Triticale 0 0 61.1 Ryegrass 100 91.9 38.9 Oat - 4.1 0 Weeds 0 4.1 0 Analyzed composition, % of DM DM 15.8 16.3 16.5 CP 17.7 18.3 16.3 NDF 41.7 44.5 49.6 ADF 22.7 24.1 27.6 Three 1.73 ac pastures were planted on a prepared seedbed for each treatment. Pastures were planted on November of 2010 and fertilized 28 d after planting with 51 lbs/ac of N (NH 4 NO 3 ) and 20 lbs/ac of S. A second fertilization with 51 lbs/ac of N (NH 4 NO 3 ) and 10 lb/ac of S took place 56 d after planting. 2 Seeding rate = 31 lb/ac of Lolium multiflorum Lam cv. Prine. 3 Seeding rate = 60 lb/ac of oat (Avena sativa cv. Horizon 201) plus 27 lb/ac of ryegrass (Lolium multiflorum Lam cv. Prine). 4 Seeding rate = 244 lb/ac of triticale (Triticosecale rimpau cv. Trical 324) plus 15 lb/ac of ryegrass (Lolium multiflorum Lam cv. Prine). 5 Average of three 2.7 ft 2 samples taken from each 1.73 ac pasture.
Table 2. Nutrient intake and digestibility by heifers fed winter forages, using Cr 2 O 3 or TiO 2 as indigestible marker and under two fecal sampling protocols. Treatment 1 Item Ryegrass Oat + ryegrass Triticale + Forage ryegrass SEM 2 treatment P-value Sampling Sampling protocol 3 Marker 4 marker 5 Intake 6, lb/d DM 9.55 10.74 9.68 1.58 0.85 - - - OM 8.51 9.48 8.73 1.41 0.88 - - - CP 1.69 1.96 1.56 0.26 0.60 - - - NDF 3.98 4.77 4.8 0.73 0.68 - - - ADF 1.94 2.29 2.42 0.37 0.64 - - - Digestibility, % DM 50.3 62.0 65.4 5.9 0.21 0.31 0.10 0.87 OM 51.8 64.8 67.5 5.8 0.18 0.25 0.10 0.91 CP 55.8 58.2 66.1 5.0 0.36 0.57 0.13 0.85 NDF 30.8 57.3 59.4 8.7 0.08 0.33 0.14 0.96 ADF 13.9 42.9 46.8 9.7 0.07 0.14 0.18 0.62 1 Winter forages were cut fresh every day from 1.73 acre pastures at 8:00 a.m. and offered ad libitum. 2 Pooled standard error of treatment means, n = 4 heifers/treatment. 3 Effect of fecal sample collection protocol: 2 samples per day (8:00 a.m. and 4:00 p.m.) vs.3 samples/d (8:00 a.m., 12:00 p.m., and 4:00 p.m.). 4 Effect of indigestible marker used to calculate apparent total tract digestibility: 10 g/d of each Cr 2 O 3 and TiO 2 were dosed once daily at 12:00 p.m. in two separate gelatin capsules. 5 Sampling protocol indigestible marker interaction. 6 Intake during the 5-d digestibility measurement period of the experiment. Table 3. Apparent total tract digestibility of nutrients measured using Cr 2 O 3 or TiO 2 as indigestible markers in heifers fed three winter forages. Marker 1 P-value Item Cr 2 O 3 TiO 2 SEM 2 effect forage 3 Marker Marker Digestibility, % DM 56.3 62.1 3.82 0.13 0.82 OM 58.6 64.2 3.75 0.13 0.81 CP 57.5 62.7 3.38 0.16 0.89 NDF 46.2 52.1 5.44 0.18 0.85 ADF 31.4 37.8 6.07 0.21 0.77 1 Heifers were dosed once daily with 10 g/d of both Cr 2 O 3 and TiO 2 once daily at 1200 h in two separate gelatin capsules. 2 Pooled standard error of treatment means, n = 4 heifers/treatment. 3 Effect of interaction between forage consumed (ryegrass, oat + ryegrass, or triticale + ryegrass) and marker used.