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This is to certify that the thesis entitled Daily Intake, Digestibility, and Rate of Digesta Passage in the Dik-Dik (Madoqua kirki) Fed Alfalfa Leaf and a Complete Pelleted Feed presented by David Jonathan Baer has been accepted towards fulfillment of the requirements for M.S. degree in Anima] SCience FuzJ—é>vetm<; CE? [7%{fiéézL Major professor Date gig/ug— g; /¢f7 0-7639 MS U is an Affirmative Action/Equal Opportunity Institution MSU LIBRARIES ”1—. V RETURNING MATERIALS: PIace in book drop to remove this checkout from your record. FINES will be charged if book is returned after the date stamped below. DAILY INTAKE, DIGESTIBILITY, AND RATE OF DIGESTA PASSAGE IN THE DIK-DIK (MADOQUA KIRK!) FED ALFALFA LEAF AND A COMPLETE PELLETED FEED By David Jonathan Baer A THESIS Submitted to Michigan State University in partial fulfillment of the requirements for the degree of MASTER OF SCIENCE Department of Animal Science 1987 ABSTRACT DAILY INTAKE, DIGESTIBILITY, AND RATE OF DIGESTA PASSAGE IN THE DIK—DIK (MADOQUA KIRKI) FED ALFALFA LEAF AND A COMPLETE FELLETED FEED By David Jonathan Baer Dik-dik (Madogua kirki) are small concentrate selectors. While metabolic body size is related to body .75 1.0 mass , gut capacity is related to body mass . To cope with this difference, small ruminants may rely on higher rates of digesta flow and selection of lower fiber feeds. The dik-dik's ability to digest forage fiber may be limited. Four captive dik-dik were fed alfalfa leaf (AL). a 1:1 ratio of AL:pelleted feed (PF) and PF to assess body weight (BW) changes, daily dry matter intake (DMI), digestibility of dry matter (DM). organic matter (OM), gross energy (GE), neutral detergent fiber (NDF), acid detergent fiber (ADF), hemicellulose and cellulose,.and the rate of digesta passage fitted by a two-pool model using a single dose of chromium- mordanted fiber. Bu, DMI, DM digestibility and 0M digestibility were similar across all treatments (P>O.10). GE digestibility was lower on AL than PF (P<0.025). NDF. ADF. hemicellulose and cellulose digestibilities were higher _on AL than PF (P<0.00i). Total mean retention time of digesta was longer on AL than PF. but there were no differences in transit time. ACKNOWLEDGEMENTS Many people have supported and encouraged me in my research and educational endeavors. I wish to express my thanks and gratitude to all of those who have helped me. Specifically, my graduate committee, especially Dr. Duane Ullrey, chairman, Dr. Melvin Yokoyama, Dr. Donald Straney and Dr. Olav Oftedal, who provided guidance during the development of my graduate program. Over the years. I have developed friendships with colleagues both at the Nutrition Lab of the National Zoological Park (NZP) and at Michigan State University. I wish to thank my friends for their support, encouragement and patience. Many individuals at NZP have supported my research. I would like to express my gratitude to Dr. Devra Kleiman. Assistant Director for Research, NZP, Dr. Olav Oftedal. Nutritionist, NZP, and Ms. Bess Frank, Collections Manager, NZP, for their assistance and interest. Friends of the National Zoo (FONZ) provided financial support for some of this research. Most importantly, I would like to acknowledge my parents and family for their continuing support and understanding. ii II. III. IV. VI. DAILY O O moow> RATE A. B. C. D. E. TABLE OF CONTENTS INTAKE AND APPARENT DIGESTIBILITY Introduction Materials and Methods Results Discussion List of References OF DIGESTA PASSAGE Introduction Materials and Methods Results Discussion List of References Appendix A Appendix 8 Appendix C Appendix D iii \It-‘(OJ-‘H H»- 20 20 22 26 28 35 38 40 43 Table Table Table Table Table Table Table Table Table Table Table Table 9. 10. 11. 12. LIST OF TABLES Ingredient composition of the pelleted feed Nutrient composition of alfalfa leaf and pelleted feed Daily dry matter intake and body weight relationships Apparent digestibility of alfalfa leaf and pelleted feed Body weight and daily dry matter intake of small ruminants Marker kinetics in the dik-dik gastrointestinal tract fed three diets: alfalfa leaf. alfalfa leafzpelleted feed. and pelleted feed Retention time of digesta in ruminants Computer program to calculate the best transit time (TT), A, and rate constant k with k and A held constant 2 i 1 Concentration of chromium in fecal dry matter of animal #5 fed pelleted feed Determination of k values using linear regression with different sets of samples Calculated best fit model parameters (k , TT, and A) with k1 and A1 held constant Model parameters for individual animals 10 10 11 27 33 38 40 40 a to Figure Figure Figure Figure Figure Figure Figure Figure Figure Figure Figure Figure Figure Figure Figure LIST OF FIGURES Chromium excretion curve. Animal #1 Chromium excretion curve, Animal #2 Chromium excretion curve, Animal #4 Chromium excretion curve, Animal #5 Chromium excretion curve, Mordanted, Animal #3 Chromium excretion curve, Mordanted, Animal #4 Chromium excretion curve, Mordanted, Animal #5 Chromium excretion curve, Mordanted, Animal #1 Chromium excretion curve, Mordanted, Animal #2 Chromium excretion curve, PF Mordanted, Animal #3 Chromium excretion curve, PF Mordanted, Animal #4 Chromium excretion curve, PF Mordanted, Animal #5 Chromium excretion curve, Animal #1 Chromium excretion curve, Animal #2 Chromium excretion curve, Animal #4 Diet: Diet: Diet: Diet: Diet: Diet: Diet: Diet: Diet: Diet: Diet: Diet: Diet: Diet: Diet: AL, AL, AL, AL, AL:PF, AL:PF, AL:PF, AL:PF, AL AL AL PF AL:PF PF AL:PF, AL:PF, AL:PF, PF, PF, PF, 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 Figure 16. LIST OF FIGURES (con’t.) Chromium excretion curve, Animal #5 vi Diet: PF, 58 1. DAILY INTAKE AND DIGESTIBILITY Introduction Herbivorous animals have evolved in a nutritional niche involving diets high in plant fiber but relatively low in digestible energy. The body size of animals, such as ruminants, that are primary consumers influences their metabolic rate (Kleiber, 1975), total energy requirement, feed intake and digestive efficiency (Demment & Van Soest, 1983; Van Soest, 1982). Dik-dik (Madoqua kirki) are among the smallest of true ruminants. The dik-dik’s feeding behavior, digestive physiology and morphology are consistent with their classification as concentrate selectors (Hofmann. 1973). As small concentrate selectors, the dik-dik’s ability to digest plant fiber may be limited. Metabolic rate is related nonlinearly to body size but 0.75) linearly to metabolic body size (Bukg . Since metabolic rate influences total nutrient and energy requirements (metabolic requirement (MR)), smaller animals have higher requirements per unit of body mass. However, herbivore gut .0 capacity (GC) is related linearly to body weight (swig > (longet weight gut contents (kg))=1.03210g(body weight .(kg))-O.936, r=.99, n=59, weight range=0.015-5177.6 kg) (Demment & Van Soest. 1983; Demment, 1982; Parra. 1978). 2 Therefore. smaller herbivores have a higher MR/GC ratio (Demment & Van Soest, 1983). To cope with this constraint, smaller ruminants may adapt in several ways. Since ruminants have evolved to use the end-products of fiber digestion as their major energy source, the extent of fiber digestion is an important factor in determining energy balance. Fiber digestion in ruminants is controlled. at least in part, by the time of contact between the substrate and the microbial population of the rumen. Therefore, factors that influence rate of digesta flow will influence digestion. These factors include morphology of the gut (e.g., ruminal pillars or restricted ostia), and physical characteristics of the feed (e.g., particle size). Dense papillation of the ruminoreticulum aids in quick absorption of volatile fatty acids (VFAs, the end-products of anaerobic microbial fermentation) (Hofmann, 1973). Behaviorally, selection of feeds that are relatively high in digestible energy (e.g., low in dietary fiber and high in nonstructural carbohydrates) will help maximize energy utilization. There are limited data on the diet of free-ranging dikF dik. Behavioral observations and stomach sample analyses indicate that the majority of their diet consists of leaves and stems. flowers, shoot tips and twigs. Consumption of fallen plant material (fresh litter) has also been observed. Some grasses, particularly leaf tips or seeds. are also consumed (Tinley, 1969; Simonetta. 1968; Hofmann, 1973; Hoppe gt 1L.. 1983). Unfortunately, there are no data on time spent foraging on different plant species or parts. There are also limited data available on the nutrient composition of these natural feeds. There are few species in the niche occupied by small ruminants (aw < 10 kg). The constraints of body size on feed intake, feed digestion and energy balance may have influenced exploitation of this niche by ruminants. Dry matter intake and digestibility by dik-dik have been investigated for only one diet, alfalfa leaves (Hoppe, 1977). Data on intake and digestibility for other small ruminants are also scarce (Nordin, 1978a,b; Cowan gt aL., 1976; Hoppe, 1977). Poor survival of dik-dik in captivity has been a problem in some zoos and may be related to nutritional management. The possibility that dik-dik can digest forage fiber to only a limited extent may be relevant to the problems of maintaining dik-dik in captivity. Feeding trials were conducted to quantitate dry matter intake by dik-dik and to assess their ability to utilize formulated diets that varied in the amount of alfalfa leaf. a relatively good quality forage fiber, and a complete pelleted feed. Nutritionally, this increase in the high quality forage:pelleted feed ratio increased the fiber levels of the diet and decreased the digestible energy density. Materials and Methods Four dik-diks were used for this study in a randomized complete block design (RCBD) with each animal as a block. Animals were reproductively mature (three males and one female) and captive born in zoos. All animals were injected with 1-2 mg lvormec (MSD Agvet, Merck and Co., Inc., Rahway, NJ) subcutaneously for control of internal and external parasites prior to use in this study. Based on records of previous physical examinations, there were no discernible health problems in these animals. The animals were individually housed in indoor 2.44 m2 pens at the National Zoological Park, Washington, DC. The pens were constructed of 0.64 cm plywood, and the walls were 1.22 m high. Concrete floors were covered with rubber matting to prevent animal slipping. Temperature and humidity were continuously recorded with a hygro—thermograph (Belfort Instrument Co., Baltimore, MD). Some natural lighting was available, but overhead fluorescent lights were set on a 12:12 L:D cycle, on at 0700 hr. Animals were sequentially fed three diets. Amounts of feed offered were based on individual ad libitum intakes of similar feeds during a 42-d preliminary period (a second female was also involved in the preliminary period). The diet for trial 1 consisted of a 1:1 ratio of alfalfa leaf (AL) from hay and a pelleted feed (PF) (Ziegler Bros., Gardners, PA) (Tables 1 and 2). The AL was manually removed from the stem and shaken through a 1.3 x 1.3 cm screen mesh 5 to remove pieces of stem. During trial I, the animals were offered 90% of their ad libitum intake to insure that a 1:1 AL:PF ratio was actually consumed. The diets for trials II and III were 100% AL and 100% PF, respectively. The animals were gradually switched from one diet to the next over a 3- to 7-d period. There was a 14-d adaptation period to the new diet prior to any collections for digestibility estimates. Following the adaptation period, total fecal collections were conducted for 7 days. Daily fecal samples collected were not pooled. Table 1. Ingredient composition of the pelleted feed (PF). Ingredient % of diet Corn grain 33.9 Alfalfa meal, sun-cured 22.0 Soybean meal (44% CF) 13.0 Alfalfa meal, dehy (17% CF) 10.0 Wheat middlings 10.0 Cane molasses 7 Soybean oil 1 Mono-dical phos (18% Ca, 21% P) 0. Salt . a 0. Vitamin premix ' 0 Mineral premix 0 Calcium propionate O aFormulated to provide per kg of diet: 6000 IU vitamin A, 500 IU vitamin D , 130 IU vitamin E, 2 mg menadione, 5 mg thiamin, 4 mg rigoflavin, 40 mg niacin, 6 mg pyridoxine, 0.2 mg biotin, 25 mg D-Ca-pantothenate, 3 mg folic acid, 30 mcg vitamin 812, 770 mg choline. bFormulated to provide per kg of diet: 100 mg Fe, 7 mg Cu, 80 mg Zn, 40 mg Mn, 700 mcg 1, 0.2 mg Se, 0.1 mg Co. 400 mg Mg. 6 Table 2. Nutrient composition of alfalfa leaf (AL) and pelleted feed (PF) (% of dry matter) (Mean:SEM). Nutrient AL (N=10) PF (N=3) Dry matter 92.7+0.5 92.5+0.7 Organic matter 83.3:0.5 85.5:0.7 Gross energy 4.5210.11 4.46:0.13 Crude protein 24.710.2 17.610.3 Ash 9.3:0.4 7.0:0.2 Neutral detergent fiber 35.710.4 21.6:0.4 Acid detergent fiber 22.7:0.5 16.2:0.3 Acid lignin 5.9:0.3 5.4:0.2 Hemicellulose 12.9:0.5 5.4:0.4 Cellulose 16.8:0.4 10.810.2 aKcal/g dry matter. Water was provided ad libitum in ceramic crooks. The cracks were washed, and the water was changed, daily. PF and AL were fed in separate containers to accurately quantify consumption of each feed item. Feed intake was monitored daily. Animals were fed daily at approximately 0900 hr. and uneaten feed was removed after approximately 24 hr and re-weighed. All offered and residual feed was weighed with an electronic balance to the nearest 0.1 g. Feces were collected daily using a small brush and a dustpan. Feces were weighed in pre-tared aluminum pans (25.4 cm diameter) lined with aluminum foil. During fecal collection periods. feces were dried in a forced air oven at 55 C for 24 hr. The dried feces were then re-weighed, stored in heavy duty plastic freezer bags and frozen (0 C) for subsequent analyses. Animals were weighed at the beginning of the study and 7 at the end of each trial. Individual animals were moved into plastic kennels (Model #200P, Doskocil Mfg. Co. Inc., Arlington, TX) and weighed with a mechanical beam balance. The sides and door of the kennels were covered with towels to darken the inside and to minimize agitation and excitement of the animals. Weights were not recorded until the animals were completely still in the kennel. Feed samples and the partially dried feces were ground in a Wiley mill fitted with a 2 mm screen. Feed and feces were analyzed for dry matter (DM), ash, gross energy (GE), neutral detergent fiber (NDF), acid detergent fiber (ADF) and acid lignin (ALIG). An electronic balance was used for all analytical weighing to the nearest 0.001 g. Dry matter was determined on approximately 0.500 g of subsample in an aluminum weighing dish by overnight drying in a forced air oven at 105 C. Ash was determined by igniting approximately 0.800 g of subsample in a muffle furnace (650 C) for'four hr. GE was determined by complete combustion in a Parr adiabatic oxygen bomb calorimeter (Parr Instrument Co., Moline, IL). Approximately 0.800 g of subsample was combusted for GE, and the remaining residue was titrated with 0.0725 N sodium carbonate to correct for the heat of formation of nitric acid. NDF and ADF were determined by the procedures of Goering and Van Soest (1970) as modified by Robertson (1978). Approximately 0.500 g and 1.000 g subsamples were used for the NDF and ADF procedures, respectively. Alpha-amylase (Catalog No. A1278. Sigma Chemical Co., St. Louis, MO.) was used in the NDF analysis (Robertson & Van Soest, 1977). ALIG was determined on the ADF using 72% sulfuric acid (Goering & Van Soest, 1970). Organic matter (OM) was calculated as DM-ash, cellulose (CELL) was calculated as ADF-ALIG, and hemicellulose (HEMI) was calculated as NDF-ADF. Crude protein (CP) was determined on feed samples with the Semiautomated Method (7.025) with approximately 0.100 g of subsample (Williams, 1984). All analyses were performed in duplicate. Total fecal collections permitted the use of the direct method to calculate apparent digestibility coefficients for OM, OM, GE, NDF, ADF, HEMI, CELL and ALIG (Schneider & Flatt. 1975). Hotelling’s multivariate T2 test was used to test for self-selection of different quality AL for two randomly selected times during the 42-d preliminary period. AL was fed to the five dik-dik. Following approximately 24 hr, the residual AL was collected. These residual AL samples, and subsamples of the offered AL, were ground and assayed for CP, NDF and ADF. A T2 statistic was calculated and compared with the critical value T20.10,3,4 = 54.97 (Gill, 1978). Analyses of variance for RCBD and orthogonal polynomials (linear and quadratic effects) (Gill, 1978) were performed on a microcomputer using LOTUS 1-2-3 software and the MS-DOS operating system. Mean dry matter intake and digestibility coefficients of 7-d collections were used in the statistical analyses. Results Mean ambient temperature (:SE) was 2410.3 C and mean relative humidity (18E) was 49:0.7% (N=18). The relatively consistent and controlled environmental conditions (light cycle, temperature, and relative humidity) permitted the sequential dietary treatments fed to all animals simultaneously. Any residual diet effects were confounded with animal and treatment. However, the controlled environment and use of standard feeding trial methodology (Schneider & Flatt, 1975), including adaptation periods to new diets, should have minimized or eliminated any residual effects.- Offered and residual AL in the preliminary trial were similar in nutrient composition (P>0.i). Thus, animals were not selecting different fractions of the offered AL, at least not on the basis of CP, NDF and ADF concentration. Therefore, in subsequent trials, residual AL was not analyzed for nutrient composition. Mean daily dry matter intake and mean body weights did not differ across treatments (Table 3). Mean daily dry matter intake per metabolic body size was 40.6. 34.6, and 0.75 kg respectively. These values were not statistically 36.1 g/BW for the AL, AL:PF, and PF trials, different. Mean dry matter and organic matter digestibilities were similar across all treatments (Table 4f. There was a slight (P<0.10) linear trend for organic matter digestibility to decrease as the proportion of AL increased. 10 Table 3. Daily dry matter intake (DM1) and body weight (BW) relationships (N=4). Factor Treatment mean SEM F --------------------- value AL AL:PF PF BW (kg) 4.23 4.28 4.48 0.08 1.08Er DMI (g) 117.5 102.2 110.5 3.9 1.81a DMI (x of BW) 75 2.85 2.42 2.48 0.08 2.81: DMI/Metabolic BW (g/BWEé > 40.8 34.6 36.1 1.1 2.51 aNot significant (P>0.10). Table 4. Mean apparent digestibility (%) of alfalfa leaf (AL) and pelleted feed (PF) from 7-d total fecal collections (N=4). Factor Treatment mean SEM F value AL AL:PF PF Dry matter 85.0 88.5 88.8 0.9 1.5: Organic matter 69.5 70.0 72.9 0.7 2.3b Gross energy 65.6 68.0 71.9 0.8 5.7c Neutral detergent fiber 66.5 48.3 36.5 0.7 143.2c Acid detergent fiber 56.6 46.6 37.9 1.0 27.6C Hemicellulose 80.8 53.1 32.7 1.0 200.7C Cellulose 66.6 55.9 41.5 1.0 57.8 a bNotsignificant (P>O.10). P<0.05. P<0.001. Whereas energy digestibility decreased as the proportion of AL increased, NDF, ADF, hemicellulose and cellulose digestibilities increased (Table 4). Gross energy digestibility (P<0.025) and fiber fraction digestibilities (P<0.001) showed very strong linear components. NDF 11 digestibility showed a very slight curvilinear component (P<0.10). No other changes in energy or fiber fraction digestibility had a curvilinear component. Discussion The daily dry matter intake for dik-dik in this study was slightly lower than previously reported. Hoppe (1977) found the average intake for adult dik-dik fed alfalfa leaf with similar gross energy density (4.50 kcal GE/g DM) to be 117 g of DM, and this DM intake corresponded to 3.76% of body weight. Hoppe (1977) suggested that this level of intake was high even though the GE density of the alfalfa leaf was high. Dry matter intakes of other small ruminants are presented in Table 5. Table 5. Body weight (BW) and daily dry matter intake (DMI) of small adult ruminants. Species - N BW DMI DMI Diet (kg) (g) (% of BW) Tragulus javanicusa 8 1.4 33.4 2.4 Mixed (hay & (Lesser mousedeer) produce) Nesotragus mochatusb 5 3.21 111 3.5 Alfalfa leaf (Suni) Cephalus monticolac 3 4 81 2.0 Alfalfa leaf (Blue duiker) (fresh) aNordin, 1978. CHoppe, 1977. Cowan gt aL., 1976. 12 Evidence from larger, domesticated ruminants supports the hypothesis that dry matter intake is controlled by gut fill and digestible energy (DE) density (kcal/g) of the diet. Gut fill is determined by volume, not weight, of the feed. Therefore, bulkiness of a diet is an important component in regulating dry matter intake. The mechanisms by which DE density regulates dry matter intake are not understood. It is possible DE density influences intake through several interrelated mechanisms in ruminants with signals from different metabolic paths (Baile and Forbes, 1974; NRC, 1987). Regardless of the mechanism, at low DE densities, dry matter intake is probably limited by gut fill (distention) and factors influencing palatability. As DE density increases, dry matter intake will increase but is still limited by gut fill and distention of the GIT. As DE density increases further, dry matter intake decreases but DE intake (kcal/d) remains constant owing to the higher DE density of the diet (Montgomery and Baumgardt, 1965; Ammann _£ 1L., 1973). Differences in DE are related to the level of NDF, a constituent of forage that is fermented slower than the cell soluble fraction. As the NDF level of a diet increases, DE and dry matter intake should decrease (Mertens, 1973). No statistical differences were found in dry matter intake across treatments for the dik-dik in this study. The current model for dry matter intake would predict that as the proportion of AL increases, and hence bulkiness and NDF level, dry matter intake would decrease. Although GE 13 densities (kcal/g) of the diets were similar, DE intake (kcal/d) was lower as the proportion of AL increased. The GE of AL in this study was similar in digestibility to that reported for comparable diets in other studies (65.6% in the current study versus 69.1% for similar quality alfalfa leaf) (Hoppe, 1977). All diets contained sufficient amounts of DE to at least meet maintenance energy requirements, as indicated by maintenance of body weight. However, dry matter intake was expected to decrease as the ratio of AL to PF increased. At steady state conditions, influx and efflux are equivalent. The amount of dry matter consumed would then be equivalent to the amount of dry matter leaving the rumen through one of two processes, passage to the lower GIT or digestion (Van Soest, 1982). The PF should have higher rates of passage than AL, related to the physical form of the PF and higher rates of digestion due to its composition and ground physical form. Why dry matter intake did not conform to current dogma is not clear. Small ruminants seem to select feeds that are readily fermented. High rates of fermentation have been reported from in vitro studies of rumen contents from free-ranging dideik. Production of between 395 and 526 umoles of fermentation gases/hr/g dry matter by dik-dik rumen contents is higher than for other East African wild ruminants studied (Clemens gt gt., 1983; Hoppe gt gt., 1983). These high fermentation rates are consistent with the relatively large 14 salivary glands of dik-dik (Hofmann, 1973; Kay gt gt., 1980) that produce buffers essential for the rumen ecosystem. Dry matter and energy digestibilities were similar to previously published data. Hoppe (1977) suggested that dry matter digestibility (67.5%) of alfalfa leaf fed to dik-dik was moderate considering the high nutritional quality of the alfalfa leaf. Organic matter digestibility has been estimated (by regression of percentage of nitrogen in the organic matter of the feces) for free-ranging dik-dik and found to be relatively high (78% and 85% for two animals) (Hofmann & Musangi, 1973). However, the regression equation used to estimate dry matter digestibility in these free- 'ranging animals was developed for sheep and may be invalid (Hoppe, 1977). Previously reported fiber digestibility coefficients for Similar diets fed to dik-dik were lower than those in the current study (54.2% for crude fiber and 39.7% for cellulose) (Hoppe, 1977). There could be several explanations for these differences. Animals in the current study were all captive born and raised whereas animals in the previous study were in captivity for only three months prior to the start of those studies. Therefore, animals in the current study may have been better adapted to alfalfa leaf than the wild caught animals. While it is difficult to make direct comparisons of the crude fiber and the detergent systems for forage fiber analysis, crude fiber best represents the ADF fraction of the detergent system (Van Soest, 1982). The crude fiber concentration in the dry 15 matter of the alfalfa leaf fed dik—dik by Hoppe (1977) was 20.9%, similar to the ADF level of 22.7% in the dry matter. of the alfalfa leaf used in the current study. Lignin is refractory to microbial fermentation and reduces fiber digestibility. The lignin content was not reported for the alfalfa fed to Hoppe’s dik-dik and was 5.9% in the current study. Methodological differences in cellulose determination may explain the large differences in cellulose digestibility. Age differences can probably be ruled out since animals were adults or reproductively mature in both studies. Fiber fraction digestibilities in this study increased in a linear fashion as the proportion of AL increased. Conversely, since dry matter digestibility was similar across treatments, the cell soluble fraction digestibility decreased as the AL increased. The cell soluble fraction was higher in the PF and was digested (either fermented in the rumen or passed to the lower tract) more readily than the NDF fraction. Thus, as the ratio of AL to PF and consequently the ratio of NDF to cell solubles increased. the animal (presumably through changes in the microbial population of the rumen) became more efficient at digesting fiber. The relative fraction of energy derived from fiber appeared to increase with the increasing AL. Similar changes in fiber fraction digestibilities have been reported in dairy cows (Uden, 1984) and horses (Thompson gt gt.. 1984) fed increasing ratios of hay to concentrate. 16 While digestion models suggest that small rUminants may be limited in their ability to digest forage fdber, the dik- dik in this study were able to extract enough energy from AL to maintain body weight. Unpublished necropsy reports from the National Zoo Department of Pathology indicated that body fat of necropsied dik-dik was low, especially during the winter months and early spring. Hoppe (1984) reported in published conference proceedings that the most fat found on one dik-dik was only 2.3% of bodyweight. Unfortunately, the methods used to measure body fat were not reported in the proceedings. These observations on body composition and energy stores suggest that dik-dik may be limited in their ability to survive prolonged cold stress. L I ST OF REFERENCES LIST OF REFERENCES Ammann, A.P., Cowan, R.L., Motherhead, C.L., & Baumgardt, B.R. (1973). Dry matter and energy intake in relation to digestibility in white-tailed deer. i.’Wildl. Lia—nasa- 37:195-201. and regulation of energy balance in ruminants. Physiol. Reviews 54:160-214. Clemens, E.T., Maloiy, G.M.0., & Sutton, J.D. (1983). Molar proportions of volatile fatty acids in the gastrointestinal tract of East African wild ruminants. Comp. Biochem. Physiol. 76A:217-224. Baile, C.A. & Forbes, J.M. (1974). Control of feed intake l Cowan, R.L., von Ketehodt, H.F. & Liebenberg, L.H.P. (1976). The blue duiker tested as a laboratory mini-ruminant. i, Anim. Sci. 43:317 (abst.). Demment, M.W. (1982). The scaling of ruminoreticulum size with body weight in East African ungulates. Afr. i. Ecol. 20:43-46. Demment, M.W. & Van Soest, P.J. (1983). Body size, digestive capacity, and fggding strategies of hggbivorgg. Morrilton, Arkansas: Winrock International. Gill, J.L. (1978). Design and analysis of experimentggin the animal and medical sciences. Ames, Iowa: The University of Iowa Press. Goering, H.K. & Van Soest, P.J. (1975). Forage fiber analyses: apparatus, reagents, procedures and some applications. Agriculture Handbook 8. Washington DC: USDA. Hofmann, R.R. (1973). The ruminant stomach: stomach structure and feeding habitg of Eggt African game ruminants. Nairobi: East African Literature Bureau. Hofmann, R.R. & Musangi, R.S. (1973). Comparative digestibility coefficients of domestic and game ruminants from marginal land in East Africa. Bull. Epi . Dis. Africa 21:385-388. 17 18 Hoppe, P.P. (1984). The physiology of the dikdik. In flgrbivore nutrition in the subtropics and tropicg:719- 722. Gilchrist, F.M.C. & Mackie, R.I. (Eds). Craighall, South Africa: The Science Press. Hoppe, P.P. (1977). Comparison of voluntary food and water consumption and digestion in Kirk’s dikdik and suni. E. Afr. Wildl. i. 15:41-48. Hoppe, P.P., van Hoven, W., von Engelhardt, Prins, R.A., Lankhorst, A., & Gwynne, M.D. (1983). Pregastric and caecal fermentation in dikdik (Madogug kirki) and suni (Nesotragus moschatus). Comp. Biochem. Physiol. 75A:517-524. Kay, R.N.B., von Engelhardt, W., & White, R.G. (1980) The digestive physiology of wild ruminants. In Digegtive physiology and metabolism in ruminants:743-761. Ruckebusch, Y.-& Thivend, P. (Eds). Westport, Connecticut: AVI Publishing Co., Inc. Kleiber, M. (1975). The firg of life:ggn introduction to animal energgticg. Huntington, New York: Kreiger. Mertens, D.R. (1973). Application of theoretical mathematical models to cell wall digestion and forage intake in ruminants. Ph.D. Thesis. Cornell University, Ithaca, New York. Montgomery, M.J. & Baumgardt, B.R. (1965). Regulation of food intake in ruminants. 1. Pelleted rations varying in energy concentration. i. Dairy Sci. 48:569-574. National Research Council. (1987). Predicting feed intgke of food-producing animals. National Academy Press: Washington DC. Nordin, M. (1978a). Voluntary food intake and digestion by the lesser mousedeer. it Wildl. Manage. 42:185-187. Nordin, M. (1978b). Nutritional physiology and behaviour of the lesser mousedeer. Biotrop. Spec. Bull. 8:83-91. Parra, R.R. (1978). Comparison of foregut and hindgut fermentation in herbivores. In Theggcology of arboregt_ folivores:205-229. Montgomery, G.G. (Ed). Washington DC: Smithsonian Institution. Robertson, J.B. (1978). The detergent system of fiber analysis. In Topicg in dietgry fiber regearch:1-42. Spiller, G.A. (Ed). New York: Plenum Press. 19 Robertson, J.B. & Van Soest, P.J. (1977). Dietary fiber estimation in concentrate feedstuffs. Proceedings of the Amer. Assoc. of Anim. Sci. Annual Meetings, University of Wisconsin, Madison, Wisconsin. p. 254. Schneider, B.H. & Flatt, W.P. (1975). The evaluation of feeds through digestibility experiments. Athens, Georgia: University of Georgia Press. Simonetta, A.M. (1966). Osservazioni etologiche ed ecologiche sui dik-dik (gen. Madogua: mammalia, bovidae) in Somalia. Monitopg Zoologico ltaligpg 75 (Suppl->zi-34. Tinley, K.L. (1969). Dikdik Madogua kirki in South West Africa: notes on distribution, ecology and behaviour. baggage 1:7-33. Thompson, K.N., Jackson, S.G. & Baker, J.P. (1984). Apparent digestion coefficients and associative effects of varying hay:grain ratios fed to horses. Nutr. Reports Intl. 30:189-197. Uden, P. (1984). Digestibility and digesta retention in dairy cows receiving hay or silage at varying concentrate levels. Anim. Feed Technology 11:279-291. Van Soest, P.J. (1964). Symposium on nutrition and forage and pastures: new chemical procedures for evaluating forages. i. Anim. Sci. 23:838-845. Van Soest, P.J. (1982). Nutritional ecology of the ruminant. Corvallis, Orgeon: 0&8 Books. Williams, S. (1984). Official methods of analysis. Arlington, Virginia: Association of Official Analytical Chemists, Inc. II. RATE OF DIGESTA PASSAGE Introduction Herbivorous animals are adapted to use the end products of anaerobic fermentation, the volatile fatty acids, as a source of energy and as a substrate for the synthesis of glucose, amino acids, and proteins. The extent of fermentation in ruminants is a function of the amount of time that feeds are in Contact with the symbiotic microbial populations of the host and the composition of the diet. Many factors have been identified that influence digesta flow through the ruminant gastrointestinal tract (GIT). Some factors are related to the physical and chemical properties of the consumed diet. These properties include particle size (Ellis et al., 1979), density (Ehle gt gt., 1984), and the rate of digestion of the feed (Mertens, 1977). Body size, the morphology of the GIT of the animal (Martens, 1973; Demment & Van Soest, 1983; Hofmann, 1973). and the level of dry matter intake, an interaction between the diet and the animal, also influence digesta flow. The terminology associated with digesta flow includes rate of digesta flow, transit time, mean retention time, turnover rate and turnover time. The rate of digesta flow is the weight or proportion of digesta that moves a specified distance in a given unit of time. The rate of 20 21 digesta flow is difficult to measure directly. However, by marking the beginning of a meal, it is possible to measure the time required for a given meal to pass through the gastrointestinal tract, which is defined as the transit time (TT). It is also possible to calculate the total mean retention time (TMRT) which is the sum of the transit time ~and the turnover times of the individual compartments of the gastrointestinal tract (Kotb and Luckey, 1972). Digesta flow through the ruminant digesta tract is a complex process. The relatively capacious fermentation vat allows for digesta mixing. The process has been mathematically modelled as a two-pool, nonreturnable system by the relationship: -k (T-TT) -k (T-TT) Y A e 1 - Ae 2 for TZTT: Y 0 for TO.1 within row. P<0.09 within row. 28 The rate constant k2 (1/hr) was statistically lower for AL than PF during the AL:PF mixed dietary treatment (P<0.08). However, no statistical difference was detected for AL and PF when they were fed individually. No statistical differences were revealed between AL and PF daring the mixed AL:PF treatment or AL and PF when fed separately for the turnover rate (hr) or for the half-life (hr) of k2. The number of turnovers/day was lower for the AL than the PF during the mixed treatment (P<0.08) but was not statistically different for the AL and PF when fed individually (Table 6). Statistical differences were not found for either of the two contrasts for the transit time (hr). AL and PF during the mixed dietary treatment had statistically similar total mean retention times (hr). However, when fed separately, AL had a statistically longer total mean retention time than PF (P<0.03) (Table 6). Discussion The major sites of fermentation are the ruminoreticulum and the cecum and colon. The development of anatomical features that delay digesta in these organs varies phylogenetically. In the rumen, some species lack morphological features such as blind sacs that trap feed, and pillars that impair feed movement. Digesta in these species should flow unimpeded. Other species which have these morphological features should have the ability to 29 retain feeds for more extensive fermentation. Concentrate selectors have the morphological characteristics commensurate with quick, unimpeded digesta flow, whereas roughage selectors have morphological characteristics that would slow the rate of digesta flow and allow fermentation of more refractory fiber fractions. Dik-dik lack the morphological features in the gastrointestinal tract (GIT) that assist in decreasing rate of digesta flow (Hofmann, 1973). The dik-dik’s ruminoreticulum is simple and S-shaped with few, poorly developed ruminal pillars. Digesta flow within the foregut is unimpeded by a wide ostium intraruminale and ostium reticulo-omasicum (Hofmann, 1973). To be effective in measuring the flow of digesta, a particulate phase marker must meet several criteria (Kotb and Luckey, 1972). Chromium-mordanted fiber fulfills many: of these criteria. The Cr in Cr-mordanted fiber is tightly bound to the NDF and is inert and non-toxic to the animal. Tight binding insures that the marker will not migrate to other digesta fractions.. In vitro and at physiological pH, the Cr is 98% recoverable from the NDF (Uden gt_gt., 1980). “The Cr-mordanted fiber can also be fed as a small proportion of the total dry matter consumed (Uden (D d gt., 1980), and_ the Cr is easily measured using atomic emission spectroscopy. The Cr is not metabolized nor absorbed from the GIT (Uden gt gt., 1980). Two criteria of Cr-mordanted fiber as a marker that remain untested are the effect of Cr-mordanted fiber on the 30 normal microflora of the GIT and the effect of Cr-mordanted fiber on the normal digestive processes. Since the Cr is tightly bound to the fiber, it is unlikely that the mordanted fiber will adversely affect the normal flora or the normal digestive processes. Another criterion for an adequate marker is that it be uniformly mixed and distributed with the digesta throughout the GIT. In an animal with rapid turnover rates, the extent to which the marker can mix with the digesta may be limited. One advantage of Cr-mordanted fiber as a marker is its likeness to the physico-chemical properties of normally consumed dietary fiber. Therefore, the Cr-mordanted fiber will flow with a digesta fraction that is biologically related to the animal, unlike some other particulate markers previously used, such as beads or chromic oxide, which flow independent of either the fluid or particulate fractions (Van Soest, 1982). Mordanting allows marking of a fraction of the actual diet. Unfortunately, the mordanting process can increase the density of the NDF. Denser particles tend to have turnover rates that are slightly higher than less dense particles (Ehle gt gt., 1984). While the density of the Cr-mordanted AL and PF were not measured in this study, different densities may be a possible source of error and variation, as the Cr concentrations of the marked NDF were certainly different. The patterns of fecal Cr excretion of the dik-dik in this study seem to fit the model. However, the 31 identification and interpretation of what the two pools represent is debated. All organs of the GIT can represent at least one pool (Stevens, 1978). Grovum and'Williams (1973) suggested that the two pools represent the turnover of digesta in the pregastric (ruminoreticulum) and postgastric (cecum and or colon) compartments. Hungate (1966) and Ellis gt gt. (1979) suggested that the two rate constants represent two pools of different sized particles in the ruminoreticulum. In any case, the declining phase represents the pool with the slowest turnover rate. The other pool has the faster turnover rate and is usually smaller. However, if turnover rates of different pools are nearly equal, it is impossible to mathematically separate the pools. There are at least three fates for a feed particle upon entering the ruminoreticulum. The feed particle can undergo digestion in the ruminoreticulum, the feed particle can be passed out of the ruminoreticulum or the feed particle can be ruminated. Mastication during the rumination process physically reduces the particle size which may increase the rate of digestion by increasing the ratio of surface area to volume of the particle as well as to increase the rate of passage. Smaller particles have higher rates of passage than larger particles. Rate of passage refers to the passage of undigested digesta. On the other hand, rumen turnover is the sum of two competing processes, the rate of digestion and the rate 32 of passage (Van Soest, 1982). Rate of digestion depends on the microbial populations and the nutrient density of the feed particle, especially the fiber and lignin content. Anything that reduces microbial growth (e.g., nitrogen deficiency, inappropriate pH) may reduce fermentation (Hungate, 1966). Rate of passage depends on the size and density of the particle, the level of dry matter intake and the morphology and body size of the host. Mean retention time of some ruminants is reported in Table 7. The two methods typically used to calculate mean retention time (Hartnell, 1977; Castle, 1956) provide similar results (Hartnell, 1977). The dik-dik in this study had overall rates of digesta retention that were slightly longer than those of other small ruminants such as suni (Table 7). While it was expected that the lesser mousedeer may have the shortest mean retention time due to its small body size, the mean retention time is actually quite long (52 hr). One explanation for this long mean retention time in the lesser mousedeer is the use of stained sorghum grain, a very novel and unusual feed item in the normal lesser mousedeer diet. Mean retention time in lesser mousedeer measured with chromium sesquioxide yielded mean retention times of 26.5 hr (Morat & Nordin, 1978). Red deer may be expected to have longer mean retention times than that reported in Table 7 because of their large size. However, the red deer used in that study were only 1 year old (Milne t al — —. ’ 1978). 33 Table 7. Mean retention time of digesta in ruminants. Species Diet Mean retention Source time (hr) Suni Alfalfa leaves 16.8 Hoppe & Gwynnwe,~1978 Suni Alfalfa stems 20.5 Hoppe & Gwynnwe, 1978 Red deer Dried grass, 21.9 Milne gt gt., 1978 pellets Dik-dik Pelleted diet 22.1 This study White tailed-deer Natural diet 22.5 Mautz & Petrides, 1971 White tailed-deer Hay, concentrates 22.7 Mautz & Petrides, 1971 a Lesser mousedeer Peanuts,lpomea 26.5 Morat & Nordin, (using Cr) 1978 Sheep Dried grass, 31.4 Milne gt gt., 1978 pellets Dik-dik Alfalfa leaf, 36.7 This study pelleted diet Goat Hay, concentrate 38.0 Castle, 1956 Dik-dik Alfalfa leaves 39.4 This study Dik-dik Alfalfa leaf, 40.9 This study pelleted diet Lesser mousedeer Peanuts,lpomea 52.8 Morat & Nordin, (using sorghum) 1978 Yak Mixed alfalfa- 78 Schaefer, 1978 brome Bison Mixed alfalfa- 79 Schaefer, 1978 brome From an energetic and morphological point of view, the retention of AL for a longer time than the PF seems contradictory. However, the physical grinding and pelleting of the PF certainly plays a part in the shorter retention time for PF than for AL. account for some differences in rate of passage. are ground tend to have faster rates of passage. The physical form of the diet may Diets that Pelleted feeds may disintegrate in the rumen and have flow 34 characteristics and rates similar to the liquid phase (Regelin, 1981). While there were no statistically significant treatment effects for the transit time, there were statistical differences in the rate constant for k1. The overall result was a statistical difference in the TMRT (hr). Contrasts between the AL and PF when fed separately indicated statistical differences in k1 while contrasts between the AL and PF when fed together revealed statistical differences in k2. These known differences limit extrapolation of information regarding rate constants of pools and digesta flow from the effects of one diet to another. Individual animals have also been identified as a major source of variation in transit time (Castle, 1956). A possible digestion strategy for dik-dik may include the selection of feeds low in fiber and relatively high in digestible energy, and an intake and rumination pattern that is constant throughout the day and rapid passage of digesta to the lower tract. Another possible adaptation to compensate for a large difference between the energy required for the metabolic body size and the energy provided for a given gut capacity may include an adaptively lower metabolic rate. Although experimental evidence of lower metabolic rates in dik-dik is limited, the data collected to 'date support this hypothesis (Kamau & Maloiy, 1981). LIST OF REFERENCES LIST OF REFERENCES Brandt, C.S. & Thacker, E.J. (1958). A concept of rate of food passage through the gastro-intestinal tract. i. Anim. Sci. 17:218-223. Castle, E. (1956). The rate of passage of foodstuffs through the alimentary tract of the goat. 1. Studies on adult animals fed hay and concentrates. Brit. ;, Nutr. 10:15- 23. Demment, M.W. & Van Soest, P.J. (1983). Body 812 L digegtive capacity, and feeding strategieg of hgrbivoreg. Morrilton, Arkansas: Winrock International. Ehle, F.R., Bas, F., Barno, B., Martin, R. & Leone, F. (1984). Particulate rumen turnover rate measurement as influenced by density of passage marker. i, Dairy Sgt. 67:2910-2913. Ellis, W.C., Matis, J.H., & Lascano, C. (1979). Quantitating rumen turnover. Fed. Proc. 38:2702-2706. Fenton, T.W. & Fenton, M. (1979). An improved procedure for to determination of chromic oxide in feed and feces. Can. ;, Anim. Sci. 59:631-634. Gill, J.L. (1978). Design gnd analysis of experiment; in the animal and medical sciences. Ames, Iowa: The University of Iowa Press. Grovum, W.L. & Williams, V.J. (1973). Rate of passage of digesta in sheep. 4. Passage of marker through the alimentary tract and the biological relevance of rate- constants derived from the changes in concentration of marker in the faeces. Brit. i. Nutr. 30:313-329. Hartnell, G.F. (1977). Measurement and significance of ingesta turnover rates in dairy cattle using rare-earth elements. Ph.D. Thesis. University of Wisconsin, Madison, Wisconsin. Hartnell, G.F. & Satter, L.D. (1979). Determination of rumen fill, retention time and ruminal turnover rates of ingesta at different stages of lactation in dairy cows. i. Anim. Sgt, 48:381-392. 35 36 Hofmann, R.R. (1973). The ruminant stomach: stomach structure and feeding hgbitg of Eggt Africgp game ruminants. Nairobi: East African Literature Bureau. Hoppe, P.P. & Gwynne, M.D. (1978). Food retention time in the digestive tract of the suni antelope (Nesotrggus moschatus). Sau etierkund. fittt. 3,235-237. Hungate, R.E. (1966). The rupenggnd its microbes. New York, New York: Academic Press. Kamau, J.M.Z. & Maloiy, G.M.O. (1981). The fasting metabolism of a small East African antelope, the dik- dik. i. Physiol. (London) 50P:319-320, Kotb, A.R. & Luckey, T.D. (1972). Markers in nutrition. Nutr. Abstr. Reviews 42:813-846. Mautz, W.W. & Petrides, G.A. (1971). Food passage rate in the white-tailed deer. i. Wildl. Manage. 35:723-731. Mertens, D.R. (1973). Application of theoretical mathematical models to cell wall digestion and forage intake in ruminants. Ph.D. Thesis. Cornell University, Ithaca, New York. Milne, J.A., Macrae, J.C., Spence, A.M., & Wilson, 5. (1978). A comparison of the voluntary intake and digestion of forages at different times of the year by the sheep and the red deer (Cervus elaphus). Brit. L. Nutr. 40:347-357. Morat, P. & Nordin, M. (1978). Maximum food intake and passage of markers in the alimentary tract of the lesser mousedeer. Mal. App . Biol. 7: 11-17. Regelin, W.L., Schwartz, C.C., Franzmann, A.W. (1981). Energy expenditure of moose on the Kenai National Wildlife Refuge. Kenai Field Station Annual Report, Kenai, Alaska. ' SAS. (1982). A user’s guide: statistics. Raleigh. North Carolina: SAS Institute, Inc. Schaefer, A.L., Young, B.A., & Chimwano, A.M. (1978). Ration digestion and retention times of digesta in domestic cattle (ggg taurus), American bison (Bison bison), and Tibetan yak (Egg grunniens). ggp, i. 2001. 56:2355- 2358. Stevens, C.E. (1978). Physiological implications of microbial digestion in the large intestine of mammals: relation to dietary factors. Amer. i. Clin. Nutr. 31:5161-5168. ' 37 Uden, P., Colucci, P.E. & Van So Investigation of chromium, in digesta. Rate of passage Agric. 31:625-632. est, P.J. (1980). cerium and cobalt as markers studies. i, Sci. Food Van Soest, P.J. (1982). Nutritiongl ecology of the ruminant. Corvallis, Orgeon: 0&8 Book 5. l III. Appendix A 38 Table 8. Computer program to calculate the best transit time (TT), A, and rate constant k2 with k1 and A1 held constant. C Metacommands for MS FORTRAN SSTORAGE:2 sNOFLOATCALLS $NODEBUG INTERFACE T0 SUBROUTINE TIME (N,STR) CHARACTER¥10 STR [NEAR,REFERENCE] INTEGER*2 N [VALUE] END INTERFACE TO SUBROUTINE DATE (N,STR) CHARACTER*1O STR [NEAR,REFERENCEJ INTEGER§2 N [VALUE] END Program for determining the best TT and R2 for the rate of passage curves fitting the data to a double exponential curve of Grovums’ method modified by Gary F. Hartnell and by David J. Baer, November, 1986 to run under MS-DOS MS-FORTRAN with an 8087 chip rectal grab samples DIMENSION T(30), P(30), A(30), PR(30) REAL*8 LIMIT,A1,STARTIME,T,A,P,PR,TT,D,R1,R2,C2, CE,81,SS,B,G,H INTEGER NUMTIME,AA,NUM CHARACTER FNAME*12, FNAME2*12, TSTR*10, DSTR*10 OPEN (5,FILE=’TEMP.DAT’) REWIND 5 50 READ (5,47) FNAME,FNAME2 READ (5,6) R1, C2, E, Bi, NUM LIMIT = 3.0 NUMTIME = 300 STARTIME = 5. OPEN (6,FILE=FNAME,STATUS='NEW’) OPEN (7,FILE=FNAME2,STATUS=’NEW’) WRITE (6,*) ’FILE: ’,FNAME WRITE (*,*) ’FILE: ’,FNAME CALL TIME (10,TSTR) WRITE (6,*) ’TIME: ’,TSTR WRITE (*,*) ’TIME: ’,TSTR CALL DATE (10,DSTR) WRITE (6,*) ’DATE: ’,DSTR C R1=K1, C2=approx. K2, E=the last time of excretion, C Bi=A1 DO 55, AA = 1,30 T(AA)=0.0 55 A(AA)=0.0 DO 56 I=1,NUM READ (5,8) T(l),A(I) 56 CONTINUE 0000000 39 Table 8 (con't.). 47 FORMAT (T1,A12,1X,A12) 6 FORMAT (2(F7.5,1X),F6.2,1X,F7.1,1X,13) 8 FORMAT (F6.2,1X,F6.3) IF (RI-99.998) 51,52,52 51 TT=STARTIME 45 R2=C2-LIMIT DO 40, N2 = 1,NUMTIME R2=R2+(LIMIT/NUMTIME) SS=O. I=O A1=EXPCDLOG(81)-R1*TT) 9 I=I+1 K=I IF (T(I)-TT) 10,10,11 10 P(I)=O. GOTO 60 11 P(I)=DLOG(A1*EXP(-R1*(T(I)-TT))-(A1*EXP(-R2*(T(I) C -TT)))) C CHI SQUARE = (OBSERVED-EXPECTED)**2/EXPECTED B=SS SS=((A(I)-P(I))**2)/ABS(P(I))+B 60 IF (T(I)-E) 9.12.12 12 IF (NZ-1) 80,80,78 78 IF (SS-D) 80,80,82 8O D=SS G=R2 H=TT DO 81, I=1,K 81 PR(I)=P(I) 82 IF (NZ-NUMTIME) 40,13,13 40 CONTINUE 13 WRITE (6,29) WRITE (6,30) D,H,R1,G,T(1),PR(1),A(1) WRITE (6,31) (T(I), PR(I), A(I), I=2,K) WRITE(7,32) D,H,R1,G 29 FORMAT (3X,’ChiSqr’,5x,’TT’,6X,’K1’,8X,’K2’,6X,’T(I)’, c2X,’PR(I)’,3X,’A(I)’) 30 FORMAT (’ ’,F8.6,3X,F5.2,3X,F6.5,3X,F7.5,2X,F6.2, 02X,F5.3,2X,F5.3) 31 FORMAT (37(38X,F6.2,2X,F5.3,2X,F5.3/)) 32 FORMAT (’ ’,F8.6,1X,F5.2,1X,F6.5,1X,F7.5) TT=TT+.25 If (TT-12.) 45,45,50 52 END IV. Appendix B 40 Table 9. Concentration of chromium (Cr) in fecal dry matter of animal #5 fed pelleted feed (PF). Sample Sampling Cr in fecal DM number time -------------------- (hr) (ppm) (In(ppm)) 1 6.13 1.00 0.000 2 7.13 1.00 0.000 3 10.13 365.40 5.901 4 12.13 894.26 6.796 5 16.13 948.61 6.855 6 18.13 970.68 6.878 7 19.13 813.22 6.701 8 21.63 707.69 6.562 9 28.63 684.71 6.529 10 32.13 434.41 6.074 11 35.63 443.63 6.095 12 36.13 348.97' 5.855 13 42.63 280.34 5.636 14 45.63 195.59 5.276 15 48.13 134.56 4.902 16 53.63 102.41 4.629 Table 10. Determination of k values using linear regression with the three sets of data paints that provide the three highest r . Set # Set of r2 In(A ) A k samples (In(ppm)) (ppm) (1/5r) used 1 6-16 0.95 8.016 3027.8 0.0604 2 7-16 0.94 8.044 3115.1 0.0610 3 6-14 0.94 7.787 2409.5 0.0518 41 Table 11. Calculated best fit model parameters (k2, TT, and A) with k and A held constant. 1 1 Set # k k TT A Chi-square (1/5r) (1/5r) (hr) (ppm) 1 0.0604 0.3504 9.25 1731.8 0.0439a 2 0.0610 0.3410 9.25 1771.2 0.0429 3 0.0518 0.5118 9.50 1473.0 0.0795 aOverall lowest chi-square. V. Appendix C 42 Table 12. Model parameters for individual animals. Diet Animal TTe A k f r2 of k f x28 # (hr) (ppm) (1}hr) k1 (1/5r) AL8 1 11.00 1903.7 0.049 0.94 0.339 0.4184 AL 2 7.00 1826.1 0.069 0.96 0.319 0.0547 AL 4 11.75 1245.7 0.033 0.92 0.233 0.2206 AL b 5 10.25 834.2 0.020 0.89 2.880 0.0488 AL:(PF) 3 7.50 1816.3 0.040 0.95 0.230 0.0948 AL:(PF) 4 11.75 1165.1 0.023 0.90 0.803 0.0752 AL:(PF) 5 11.25 2006.8 0.052 0.94 0.192 0.1459 (AL):PF 1 11.00 186.8 0.027 0.78 2.897 0.1053 (AL):PF 2 11.75 1863.6 0.057 0.96 0.557 0.0530 (AL):PF 3 11.25 1558.1 0.089 0.98 3.119 0.0465 (AL):PF 4 11.25 1015.1 0.031 0.83 3.061 0.0385 (Ah):PF 5 10.25 2168.4 0.046 0.94 0.176 0.0254 PF 1 11.00 1354.2 0.139 0.90 0.329 0.1558 PF 2 5.75 4517.0 0.155 0.99 0.315 0.8756 PF 4 11.75 1841.5 0.058 0.93 0.298 0.1202 PF 5 9.25 1771.2 0.061 0.94 0.341 0.0429 :Alfalfa leaf. cMarked alfalfa leaf - mixed diet. dMarked pelleted feed - mixed diet. ePelleted feed. fTT = transit time. k1 and k2 = rate constants. 8X2 = calculated chi-square value. VI. 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