Effects of Urea and Starch on Rumen Fermentation, Nutrient Passage to the Duodenum, and Performance of Cows' M. R. CAMERON, 1. H. KLUSMEYER, G. L. LYNCH? and J. H. CLARK3 Deparhnent of Animal Sciences D. R. NELSON Department of Veterinary Clinical Medicine University of Illinois Urbana 61801
starch. Feeding urea increased passage of methionine to the small intestine, whereas feeding additional starch increased passage of methionine and arginine. Passage of other amino acids to the small intestine was not altered significantly by feeding urea or additional starch. Production of milk and milk protein was increased, but yields of fat and SNF were not altered by feeding diets supplemented with urea. Production of milk and milk fat was not affected,but yields of CP and SNF were decreased when additional starch was fed to cows. (Key words: rumen fermentation, urea, starch, microbial protein synthesis)
ABSTRACT
Four midlactation. multimrous Holstein cows fitted with r u m i h and duodenal cannulas were used in a 4 x 4 Latin square design to determine the effects of supplementing urea or starch or both to diets containing fish meal on passage of nutrients to the small intestine and performance of lactating cows. The treatments (in a 2 x 2 factorial arrangement) were 1) control and control plus 2) urea, 3) starch, or 4) starch and urea. Supplementing diets with urea did not affect DMI; ruminal, postruminal, or total tract digestibilities of DM, starch, ADF, or NDE ruminal fluid VFA concentrations or molar percentages; or ruminal fluid or particulate dilution rates. Feeding additional starch depressed DMI but did not alter ruminal or postruminal digestion of OM or W A concentrations and molar percentages in ruminal fluid. Ruminal fluid ammonia concentration was increased by feeding urea and decreased by feeding additional starch. Passage of nonammonia N, nonammonia nonmicrobial N, or microbial N to the small intestine and efficiency of microbial CP synthesis were not a€fected significantly by supplying either urea or additional
Abbreviation key: NANMN = nonammonia nonmicrobial N, OMAD = apparently digested OM, OMTD = truly digested OM. INTRODUCTION
Synthesis of microbial protein and growth of ruminal microbes depends on adequate energy and N for synthesis and assimilation of amino
Received April 9, 1990. Accepted September 4, 1990. 'Supported in part by thc University of Illinois Agricultnral Experiment Station and a gitt from the Commcdity Credit Corporation, USDA, 2BASP corporatioq 100 Chemy Hill Rd., Parsipparry, NI 07054. 'Address unrespondcnce to: J i i H.Chk,Department of Animal Sciences, University of Illinois, 315 Animal Sciences Lab, I207 W. Oregory Drive, U r b a ~ ,IL. 61801. 1991 J Dairy SCi 741321-1336
acids. Simultaneous ruminal degradation and utilization of dietary protein and carbohydrate are necessary for optimal microbial growth and protein synthesis (36). Energy is one of the most limiting factors for microbial growth and protein synthesis in the rumen of dairy cows. Lactating cows traditionally are fed high quality forages and soybean meal containing protein that is rapidly degraded in the rumen (26). However, a shortage of N (NH3, amino acids, or peptides) also may limit microbial growth and protein synthesis in the m e n , especially when diets containing a high concentration of ruminally nondegradable protein are fed (24, 27).
1321
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CAMERON ET AL.
The current practice for increasing postruminal supply of amino acids in lactating cows is to formulate diets that contain protein supple ments with a low degradability in the m e n . Fish meal contains protein that has a low degradability m the rumen; it has been used in formulating diets for dairy cows in an attempt to increase amino acid supply to the small intestine (26). However, responses in both amino acid passage to the small intestine and milk production have not always been positive with dietary inclusion of fish meal. In the trials of Zerbini et al. (48) and McCarthy et al. (25), ruminal N H 3 concentrations were significantly lower for cows fed fish meal-supplemented diets than for those fed soybean meal-supple mented diets. Zerbini et al. (48) postulated that N H 3 may have limited microbial protein synthesis when cows were fed fish meal-supple mented diets. The low ruminal degradability of fish meal may have led to a reduction in the amount of energy available to the microbes for growth. The objective of our experiment was to investigate the effects of supplementing a diet containing fish meal with 1) urea as a source of NH3,2) starch as a source of energy, or 3) urea and starch on ruminal fermentation, nutrient passage to the small intestine, apparent total tract nutrient digestibility, and performance of lactating dairy cows.
urea. Ingredient and chemical composition of
each diet is shown in Table 1. Supplemental starch was a corn starch crumble or “grit’ (AE Staley, Decatur, IL)that replaced corn silage in the control diet. Diets were fed for ad libitum intake as a TMR twice daily at 0500 and 1700 h. Cows were housed in conventional stanchions and milked twice daily at 0500 and 1600 h. Cows were allowed to exercise daily from 0600 to 0700 h except for the last 3 d of each period. Dry Matter Intake, Milk Production, and Mllk Composition
Feed intakes were measured and m r d e d daily. Samples of concentrates, alfalfa haylage, and corn silage were obtained daily during the last 5 d of each period and composited. Feed refusals also were obtained daily from each cow during the last 5 d of each period and pooled for each cow on a pmportional basis according to the amount of feed refused each day. The samples were dried at 55’C to constant weight to determine DM content. Dry samples were ground in a Wiley mill (Arthur A. Thomas, Philadelphia, FA) through a 1-mm screen. The composited samples were analyzed for OM ( W C for 8 h), CP (2). ADF (ll), and NDF the method of Goering and Van Soest (11) as modified by Jeraci et al. (IS)]. Starch was determined by enzymatic hyMATERIALS AND METHODS drolysis (17). Cornposited samples were exposed to acid hydrolysis (10) and amino acid Cows, Treatments, and concentration was detexmined by an automated Experlmental Deslgn amino acid analyzer (Beckman 6300, Beckman Four multiparous Holstein cows surgically Instruments, Fullerton, CA). Milk weights were recorded at each milking. fitted with ruminal and duodenal cannulas prior to parturition were used in the experiment. The During the last 7 d of each period, milk samruminal cannulas were made of sofi plastic and ples were composited daily according to yield were 10.2 cm in diameter. The duodenal cannu- for both the am. and p.m. milkings, preserved las were of the T-shape design described by with 2-bromo-2-nitropropane-l,3-diol,and Komarek (21); they were placed proximal to stored at 4 C until a n a l y d for SNF (12). the common bile and pancreatic duct, approxi- protein, and fat contents (infrared analysis; mately 10 cm distal to the pylorus. The cows Dairy Lab Services, Inc., Dubuque, IA). averaged 146 d postpartum (range, 136 to 156 Rumlnal Fermentatlon and d) at the beginning of the experiment. The experimental design was a 4 x 4 Latin Nutrient Flow to square with a 2 x 2 factorial arrangement of the Small Intestine treatments. Each experimental period cmsisted Starting at 1800 h on d 11, ruminal fluid of 14 d. The four treatments were 1) control samples were collected every 3 h for the last 3 diet, 2) control diet plus urea, 3) control diet d of each period. Sampling was staggered 1 h plus starch, and 4) control diet plus starch and daily so that every hour in a 24-h day would be Journal of Dairy Science Vol. 74, No. 4, 1991
EFFECK3 OF UREA
1323
AND STARCH ON RUMEN FERMENTATION
represented. Ruminal fluid was obtained through the ruminal cannulas via a suction pump. The pH of the ruminal fluid was determined immediately using a glass electrode. Samples were acidified @H I 2) with 50% sulfuric acid and centrifuged at 27,000 x g for 10 min. The supernatant was decanted and fkoZen until analysis. After thawing, ruminal fluid samples were analyzed for NH3, VFA, and Co concentrations. Ammonia concentrations in ruminal fluid were determined by the colorimetric assay of Chaney and Marbach ( 5 ) as modified by Cotta and Russell (6). Cancentrations of W A in ruminal fluid were determined according to the pmcedures of Klusmeyer et al. (18) with an automated gas chromatograph (Model 4600, Varian, Palo Alto, CA). Ruminal fluid dilution rate, volume, and outflow were determined using Co as a marker
(44). At 2100 h on d 11 of each period, each cow was dosed with 50 g of sodium cobalt EDTA dissolved in 500 ml of water (44). A
plastic tubing fitted with a plastic funnel was pushed through the ruminal mat, and the Co solution was poured into several locations within the rumen. Ruminal fluid samples, obtained 3, 6, 9, 12, 15, 18, 22, and 25 h after dosing, were analyzed for Co content using an atomic absorption spectrophotometer (Model 2380, Perkin-Elmer Corp., Norwalk, cr).Ruminal fluid dilution rate, volume, and total outflow were calculated using regression analySiS.
At 1700 h on d 11 of each period, each cow was fed 2.5 kg of TMR sprayed with a solution of ytterbium chloride (2.5 g YbmO ml of water) to determine particulate passage rate from the rumen Ruminal digesta samples were
TABLE 1. In@dieat and chemical composition of diets as
Item Ingredient M a a hylage' corn silage2
Ground shelled corn Fish meal3 Sodium bicarbonate Sodium chloride M i m d and vitamin mix4
Dicalcium phosphate Limstone Magnesium oxide
Una
35 .00 20.00 39.40 3.00 .75 .15 .15
35.00 20.00 38.65 3.00 .75 .15 .I5 .40 .35 .15 .65 .I5
.35 .15
.a
Urea
-
percentage of DM.
Control
.40
Sodium sulfate
8
SWh
Starch +urea
35.00 7.50 37.05 5.00 .75 .15 .15 1.MI
35.00 750 36.30 5.00 .75 .15 .15 1.00
23
.23 .67 .75 9.38 3.12
.67 9.38 3.12
Starch Dextrose Chemical component DM OM
732 90.9 14.9 34.4 195 32.7
CP Starch
ADF NDF
733 902 16.9 33.9 19.4 32.6
78.8 91.0 14.8 40.6 16.2 275
78.8 90.3 16.8 40.2 162 27.4
~
l.m
'Contains: OM, 64.3% OM, 87.8%; CP. 20.7%; starch, ADF, 36.7% and NDF, 46.1%. konblins: DM, 49.4% OM, 94.6% CY. 9.3% starch. 34.096,ADF. 25.996, and NDF. 44.6%. 3Cmtains:DM, 91.1%OM,76.096, etha extract, 8.6% CP,73.9%;and soluble N,9.4%in 10% Burroughs miaeral mixture.
4ContaioS: Co, .W%; ell, 5 %I, ,02546;Fe, 2.096, Mg, 5.W;Mu, 3.0% & 75%;Se, .015% S, 10.096; Zn, 3.096, vitamin A, 2200 KJ/g vitamin q,660 JU/g and vitamin E, 8 IU/g. Jolprnal
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CAMWON ET AL.
collected 4, 10, 16, 22, 29, 35, 41, and 47 h after dosing with Yb. The digesta samples were dried at 55'C to constant weight (about 72 h), ground in a Wiley mill (1-mm screen), and analyzed for Yb concentration using an atomic absorption spectrophotometer according to procedures of Ellis et al. (7)using 3 M HNO3 and 3 M HCl solutions. Regression analysis was used to determine particulate (Yb) flow rate from the rumen. Duodenal samples were obtained at the same time as ruminal fluid samples during d 11 to 14 of each period. Approximately 400 ml of duodenal digesta from each cow far each hour of the day were collected and frozen. Upon thawing, all 24 duodenal samples from individual cows were pooled and mixed thoroughly. A p proximately 4 L of the compositsd duodenal digesta were refrozen, lyophiked, and ground in a Wiley mill (1-mm screen). The duodenal digesta samples were analyzed for OM, CP, ADF, NDF, starch, and amino acids by proce dures described. Ammonia N in the duodenal digesta was detennined by steam distiUation with magnesium oxide as described by Bremner and Keeney (3). Duodenal digesta also were analyzed for concentration of purines (49), which served as the bacterial flow marker. Digesta contents were collected from the reticulum 1, 3, 5, 7, 9, and 11 h after feeding for the isolation of bacteria Contents were collected by forcing a 500-ml bottle through the ruminal mat to the reticulo-omasal orifice, allowing it to fill with digesta, and then withdrawing the bottle. For each sample, bacteria were isolated by differential centrifugation using the method of Steinhour et al. (40). Bacteria collected from each cow were pooled within a period, frozen, and lyophilized. The resulting bacterial samples were analyzed for DM, OM, CP, and purines by the methods described. Rumlnal and Total Tract Digest1bllltles
ples were frozen at the time of collection, thawed, and composited for each cow on an equal wet weight basis at the end of each period. The composited samples were dried at 55'C and ground in a Wiley mill (1-mm screen). Dry matter, OM, CP,starch, ADF, and NDF were detennined using methods described above. Both fecal and duodenal samples were assayed for Cr content by atomic absorption spectrophotometry using procedures described by Williams et al. (45). Statistical Analysis
Data were analyzed by ANOVA for a Latin square design using the general hear model procedure of SAS (37). The single degree of freedom, orthogonal comparisons for the treatment means were 1) urea versus no urea, 2) high starch versus low starch, and 3) the interaction between urea and starch. RESULTS AND DISCUSSION
Nutrient Content
of
Diets
Diets supplemented with starch contained more DM and starch, but less ADF and NDF, than diets not supplemented with starch, because starch replaced corn silage in the diet (Table 1). Urea supplied about 12.5% of the total N in the urea-supplemented diets, and these diets contained about two percentage units more CP than diets not supplemented with urea Alfalfa haylage supplied the largest percentage (42 to 49%) of 8 in the experimental diets. Fish meal (Zapata Haynie Corp., Reedville, VA), the source of supplemental protein, supplied 14.9, 13.1,25.0,and 22.0% of the total N in the control, urea, starch, and starch plus urea diets, respectively. Intakes and Dlgestlbllltles
Efects
of Urea.
Intake, ruminal digestion,
Chromic oxide was used as a marker to and passage to the duodenum of DM, OM, measure ruminal and total tract digestibilities of starch, ADF, and NDF were not affected by DM, OM, CP, ADF, NDF, and starch. Twice supplementing urea in the diet (Tables 2 and 3). daily, at 0700 and 1900 h, on d 5 through 14 of The daily quantities (kilograms per day) of each period, 10 g of chromic oxide were placed in the rumen via the ruminal cannula. Fecal grab samples were obtained at 0700 and 1900 h during the last 5 d of each period. F e d samJouIIlal of Dairy Science VoL 74, No. 4, 1991
OM, starch, ADF, and NDF digested postnuninally also were not altered by supplementing urea in the diet, but OM digested postruminally expressed either as a percentage of OM intake
EpFE(JTs
1325
OF UREA AND STARCH ON RUMEN FERMENTATION
(P< .07)or as a percentage of OM that passed to the duodenum (P < .@ was increased I) by supplementing urea in the diet (Table 2). The significantly greater percentage of OM (62.8 vs. 57.7%) digested postruminally for diets s u p plemented with urea compared with diets not supplemented with urea can be explained by the concomitant increase in percentage of fiber digested postruminally (Table 3). Digestibilities of ADF (P < .09) and NDF (P < .11) that passed to the small intestine were increased by 5.4 and 5.7 percentage units, respectively, when urea was added to the diet (Table 3). Therefore, small increases in total tract apparent digestibility coefficients were obtained for DM, OM, starch, ADF, and NDF when urea was added to the diet, but these small increases were not significant Vables 2 and 3). Our data support previous reports (8, 46) indicating that DMI of lactating dairy cows was not affected by feeding less than 1.0% of the
total dietary DM as u r a In contrast, Polan et al. (32) depressed DMI of lactating dairy cows when urea was included in the concentrate to supply 1% of the total dietary DM. As we Observed with lactating COWS, ruminal DM digestion was not affected, but postruminal DM disappearance! was significantly improved by infusing 9.8 g/d of urea into the rumen of sheep fed high concentrate diets (31). Likewise, Ortigues et al. (30) reported that ruminal digestion of DM and ADF was not affected by supplementing urea in diets consisting of fescue hay. However, postruminal digestion of DM and ADF was improved when urea was supplemented to the diet, because the quantity of DM and ADF that disappeared from the cecum and colon was increased, whereas the amounts of DM and ADF digested in the small intestine were not altered significantly. Therefore, the i m p v e d postrumjnal digestibilities of DM, OM, ADF, and NDF may have been the result
TABLE 2. Least squares means for intake and digestibility of DM, OM, and starch in various segments of the gastrointestinal tract of lactating cows fed diets that contained fish meal and were supplemented with starch, urea, or starch plus urea Starch
Item
DM h e ,Wd ADl-r.1 96
Control Urea
Starch
+urea
SEM
Urea
Starch
23.1 69.0
23.0 712
21.6 68.0
21.0 71.3
.6 1.7
57 .14
.78
21.0 6.1 29.8 9.8 47.2 14.9 8.7 40.9 57.4 70.6
20.9 5.3 25.6 9.1 44.0 15.6 9.9 472 62.9 72.7
19.6 5.4 8.8 44.9 14.2 8.3 42.1 57.9 70.1
19.1 52 27.6 8.9 46.6 13.9 8.9 46.0 62.7 73.5
5 .7 3.0 .6 2.6 .6 .6 2.3 1.9 1.7
7.6 3.4 45.5 42 4.0 515
7.5 2.6 34.4 4.9 4.7 62.3
8.3 4.1 49.1 4.2 4.0 47.4
8.0 4.2 52.4 3.8 3.7 46.0
2 .4 4.4 A .4 4.1
94.3
94.6 96.7
92.6 96.5
95.4 985
1.3 .7
.02
OM h a ,Wd Apparently digested in the m e n , kg/d Percentage of OM intake Truly digested in the m e n , kg/d Percentage of OM intake Passage to duodenum, kg/d Appanntty digested P o ~ Wd , PCTcentage of OM mtake Percentage of passage to duodenum ADlT,1 %
28.0
58
.14
.02 59 .98 .30 .98 .ll 22 .98 .94 .94
.46 .46
.03 .03
.46 .47
.64 .78 .75 .16
.W .04
Starch
Intake, Wd Apparently digested in the rumen, kg/d Percentage of starch intake Passage to duodenum, kg/d ApP-aY diststed postnmunau ’ Y, kg/d Percentage of starch intake Percentage of starch passage to duodenum ADlT.1 %
97.0
A1
.os
.70
30
55 29
21 .05
26 29
.74 .39
%teraction bestarch and urea was not significant (P > .lo). l~pparentlydigested in thc total g~tmintestinaltract
J o d of Dairy Science Vol. 74. No. 4, 1991
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CAMERON ET AL.
TABLE 3. Least squares means for intake and digestibility of ADP and NDP in various segments of the gastrOinttstiaal tract of lactating cows fed dicts that c o n t a i d fish meal and werc supplemented with starch, urea, or starch plus una. significance of difference (P
starch. Feeding urea increased passage of methionine to the small intestine, whereas feeding additional starch increased passage of methionine and arginine. Passage of other amino acids to the small intestine was not altered significantly by feeding urea or additional starch. Production of milk and milk protein was increased, but yields of fat and SNF were not altered by feeding diets supplemented with urea. Production of milk and milk fat was not affected,but yields of CP and SNF were decreased when additional starch was fed to cows. (Key words: rumen fermentation, urea, starch, microbial protein synthesis)
ABSTRACT
Four midlactation. multimrous Holstein cows fitted with r u m i h and duodenal cannulas were used in a 4 x 4 Latin square design to determine the effects of supplementing urea or starch or both to diets containing fish meal on passage of nutrients to the small intestine and performance of lactating cows. The treatments (in a 2 x 2 factorial arrangement) were 1) control and control plus 2) urea, 3) starch, or 4) starch and urea. Supplementing diets with urea did not affect DMI; ruminal, postruminal, or total tract digestibilities of DM, starch, ADF, or NDE ruminal fluid VFA concentrations or molar percentages; or ruminal fluid or particulate dilution rates. Feeding additional starch depressed DMI but did not alter ruminal or postruminal digestion of OM or W A concentrations and molar percentages in ruminal fluid. Ruminal fluid ammonia concentration was increased by feeding urea and decreased by feeding additional starch. Passage of nonammonia N, nonammonia nonmicrobial N, or microbial N to the small intestine and efficiency of microbial CP synthesis were not a€fected significantly by supplying either urea or additional
Abbreviation key: NANMN = nonammonia nonmicrobial N, OMAD = apparently digested OM, OMTD = truly digested OM. INTRODUCTION
Synthesis of microbial protein and growth of ruminal microbes depends on adequate energy and N for synthesis and assimilation of amino
Received April 9, 1990. Accepted September 4, 1990. 'Supported in part by thc University of Illinois Agricultnral Experiment Station and a gitt from the Commcdity Credit Corporation, USDA, 2BASP corporatioq 100 Chemy Hill Rd., Parsipparry, NI 07054. 'Address unrespondcnce to: J i i H.Chk,Department of Animal Sciences, University of Illinois, 315 Animal Sciences Lab, I207 W. Oregory Drive, U r b a ~ ,IL. 61801. 1991 J Dairy SCi 741321-1336
acids. Simultaneous ruminal degradation and utilization of dietary protein and carbohydrate are necessary for optimal microbial growth and protein synthesis (36). Energy is one of the most limiting factors for microbial growth and protein synthesis in the rumen of dairy cows. Lactating cows traditionally are fed high quality forages and soybean meal containing protein that is rapidly degraded in the rumen (26). However, a shortage of N (NH3, amino acids, or peptides) also may limit microbial growth and protein synthesis in the m e n , especially when diets containing a high concentration of ruminally nondegradable protein are fed (24, 27).
1321
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CAMERON ET AL.
The current practice for increasing postruminal supply of amino acids in lactating cows is to formulate diets that contain protein supple ments with a low degradability in the m e n . Fish meal contains protein that has a low degradability m the rumen; it has been used in formulating diets for dairy cows in an attempt to increase amino acid supply to the small intestine (26). However, responses in both amino acid passage to the small intestine and milk production have not always been positive with dietary inclusion of fish meal. In the trials of Zerbini et al. (48) and McCarthy et al. (25), ruminal N H 3 concentrations were significantly lower for cows fed fish meal-supplemented diets than for those fed soybean meal-supple mented diets. Zerbini et al. (48) postulated that N H 3 may have limited microbial protein synthesis when cows were fed fish meal-supple mented diets. The low ruminal degradability of fish meal may have led to a reduction in the amount of energy available to the microbes for growth. The objective of our experiment was to investigate the effects of supplementing a diet containing fish meal with 1) urea as a source of NH3,2) starch as a source of energy, or 3) urea and starch on ruminal fermentation, nutrient passage to the small intestine, apparent total tract nutrient digestibility, and performance of lactating dairy cows.
urea. Ingredient and chemical composition of
each diet is shown in Table 1. Supplemental starch was a corn starch crumble or “grit’ (AE Staley, Decatur, IL)that replaced corn silage in the control diet. Diets were fed for ad libitum intake as a TMR twice daily at 0500 and 1700 h. Cows were housed in conventional stanchions and milked twice daily at 0500 and 1600 h. Cows were allowed to exercise daily from 0600 to 0700 h except for the last 3 d of each period. Dry Matter Intake, Milk Production, and Mllk Composition
Feed intakes were measured and m r d e d daily. Samples of concentrates, alfalfa haylage, and corn silage were obtained daily during the last 5 d of each period and composited. Feed refusals also were obtained daily from each cow during the last 5 d of each period and pooled for each cow on a pmportional basis according to the amount of feed refused each day. The samples were dried at 55’C to constant weight to determine DM content. Dry samples were ground in a Wiley mill (Arthur A. Thomas, Philadelphia, FA) through a 1-mm screen. The composited samples were analyzed for OM ( W C for 8 h), CP (2). ADF (ll), and NDF the method of Goering and Van Soest (11) as modified by Jeraci et al. (IS)]. Starch was determined by enzymatic hyMATERIALS AND METHODS drolysis (17). Cornposited samples were exposed to acid hydrolysis (10) and amino acid Cows, Treatments, and concentration was detexmined by an automated Experlmental Deslgn amino acid analyzer (Beckman 6300, Beckman Four multiparous Holstein cows surgically Instruments, Fullerton, CA). Milk weights were recorded at each milking. fitted with ruminal and duodenal cannulas prior to parturition were used in the experiment. The During the last 7 d of each period, milk samruminal cannulas were made of sofi plastic and ples were composited daily according to yield were 10.2 cm in diameter. The duodenal cannu- for both the am. and p.m. milkings, preserved las were of the T-shape design described by with 2-bromo-2-nitropropane-l,3-diol,and Komarek (21); they were placed proximal to stored at 4 C until a n a l y d for SNF (12). the common bile and pancreatic duct, approxi- protein, and fat contents (infrared analysis; mately 10 cm distal to the pylorus. The cows Dairy Lab Services, Inc., Dubuque, IA). averaged 146 d postpartum (range, 136 to 156 Rumlnal Fermentatlon and d) at the beginning of the experiment. The experimental design was a 4 x 4 Latin Nutrient Flow to square with a 2 x 2 factorial arrangement of the Small Intestine treatments. Each experimental period cmsisted Starting at 1800 h on d 11, ruminal fluid of 14 d. The four treatments were 1) control samples were collected every 3 h for the last 3 diet, 2) control diet plus urea, 3) control diet d of each period. Sampling was staggered 1 h plus starch, and 4) control diet plus starch and daily so that every hour in a 24-h day would be Journal of Dairy Science Vol. 74, No. 4, 1991
EFFECK3 OF UREA
1323
AND STARCH ON RUMEN FERMENTATION
represented. Ruminal fluid was obtained through the ruminal cannulas via a suction pump. The pH of the ruminal fluid was determined immediately using a glass electrode. Samples were acidified @H I 2) with 50% sulfuric acid and centrifuged at 27,000 x g for 10 min. The supernatant was decanted and fkoZen until analysis. After thawing, ruminal fluid samples were analyzed for NH3, VFA, and Co concentrations. Ammonia concentrations in ruminal fluid were determined by the colorimetric assay of Chaney and Marbach ( 5 ) as modified by Cotta and Russell (6). Cancentrations of W A in ruminal fluid were determined according to the pmcedures of Klusmeyer et al. (18) with an automated gas chromatograph (Model 4600, Varian, Palo Alto, CA). Ruminal fluid dilution rate, volume, and outflow were determined using Co as a marker
(44). At 2100 h on d 11 of each period, each cow was dosed with 50 g of sodium cobalt EDTA dissolved in 500 ml of water (44). A
plastic tubing fitted with a plastic funnel was pushed through the ruminal mat, and the Co solution was poured into several locations within the rumen. Ruminal fluid samples, obtained 3, 6, 9, 12, 15, 18, 22, and 25 h after dosing, were analyzed for Co content using an atomic absorption spectrophotometer (Model 2380, Perkin-Elmer Corp., Norwalk, cr).Ruminal fluid dilution rate, volume, and total outflow were calculated using regression analySiS.
At 1700 h on d 11 of each period, each cow was fed 2.5 kg of TMR sprayed with a solution of ytterbium chloride (2.5 g YbmO ml of water) to determine particulate passage rate from the rumen Ruminal digesta samples were
TABLE 1. In@dieat and chemical composition of diets as
Item Ingredient M a a hylage' corn silage2
Ground shelled corn Fish meal3 Sodium bicarbonate Sodium chloride M i m d and vitamin mix4
Dicalcium phosphate Limstone Magnesium oxide
Una
35 .00 20.00 39.40 3.00 .75 .15 .15
35.00 20.00 38.65 3.00 .75 .15 .I5 .40 .35 .15 .65 .I5
.35 .15
.a
Urea
-
percentage of DM.
Control
.40
Sodium sulfate
8
SWh
Starch +urea
35.00 7.50 37.05 5.00 .75 .15 .15 1.MI
35.00 750 36.30 5.00 .75 .15 .15 1.00
23
.23 .67 .75 9.38 3.12
.67 9.38 3.12
Starch Dextrose Chemical component DM OM
732 90.9 14.9 34.4 195 32.7
CP Starch
ADF NDF
733 902 16.9 33.9 19.4 32.6
78.8 91.0 14.8 40.6 16.2 275
78.8 90.3 16.8 40.2 162 27.4
~
l.m
'Contains: OM, 64.3% OM, 87.8%; CP. 20.7%; starch, ADF, 36.7% and NDF, 46.1%. konblins: DM, 49.4% OM, 94.6% CY. 9.3% starch. 34.096,ADF. 25.996, and NDF. 44.6%. 3Cmtains:DM, 91.1%OM,76.096, etha extract, 8.6% CP,73.9%;and soluble N,9.4%in 10% Burroughs miaeral mixture.
4ContaioS: Co, .W%; ell, 5 %I, ,02546;Fe, 2.096, Mg, 5.W;Mu, 3.0% & 75%;Se, .015% S, 10.096; Zn, 3.096, vitamin A, 2200 KJ/g vitamin q,660 JU/g and vitamin E, 8 IU/g. Jolprnal
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collected 4, 10, 16, 22, 29, 35, 41, and 47 h after dosing with Yb. The digesta samples were dried at 55'C to constant weight (about 72 h), ground in a Wiley mill (1-mm screen), and analyzed for Yb concentration using an atomic absorption spectrophotometer according to procedures of Ellis et al. (7)using 3 M HNO3 and 3 M HCl solutions. Regression analysis was used to determine particulate (Yb) flow rate from the rumen. Duodenal samples were obtained at the same time as ruminal fluid samples during d 11 to 14 of each period. Approximately 400 ml of duodenal digesta from each cow far each hour of the day were collected and frozen. Upon thawing, all 24 duodenal samples from individual cows were pooled and mixed thoroughly. A p proximately 4 L of the compositsd duodenal digesta were refrozen, lyophiked, and ground in a Wiley mill (1-mm screen). The duodenal digesta samples were analyzed for OM, CP, ADF, NDF, starch, and amino acids by proce dures described. Ammonia N in the duodenal digesta was detennined by steam distiUation with magnesium oxide as described by Bremner and Keeney (3). Duodenal digesta also were analyzed for concentration of purines (49), which served as the bacterial flow marker. Digesta contents were collected from the reticulum 1, 3, 5, 7, 9, and 11 h after feeding for the isolation of bacteria Contents were collected by forcing a 500-ml bottle through the ruminal mat to the reticulo-omasal orifice, allowing it to fill with digesta, and then withdrawing the bottle. For each sample, bacteria were isolated by differential centrifugation using the method of Steinhour et al. (40). Bacteria collected from each cow were pooled within a period, frozen, and lyophilized. The resulting bacterial samples were analyzed for DM, OM, CP, and purines by the methods described. Rumlnal and Total Tract Digest1bllltles
ples were frozen at the time of collection, thawed, and composited for each cow on an equal wet weight basis at the end of each period. The composited samples were dried at 55'C and ground in a Wiley mill (1-mm screen). Dry matter, OM, CP,starch, ADF, and NDF were detennined using methods described above. Both fecal and duodenal samples were assayed for Cr content by atomic absorption spectrophotometry using procedures described by Williams et al. (45). Statistical Analysis
Data were analyzed by ANOVA for a Latin square design using the general hear model procedure of SAS (37). The single degree of freedom, orthogonal comparisons for the treatment means were 1) urea versus no urea, 2) high starch versus low starch, and 3) the interaction between urea and starch. RESULTS AND DISCUSSION
Nutrient Content
of
Diets
Diets supplemented with starch contained more DM and starch, but less ADF and NDF, than diets not supplemented with starch, because starch replaced corn silage in the diet (Table 1). Urea supplied about 12.5% of the total N in the urea-supplemented diets, and these diets contained about two percentage units more CP than diets not supplemented with urea Alfalfa haylage supplied the largest percentage (42 to 49%) of 8 in the experimental diets. Fish meal (Zapata Haynie Corp., Reedville, VA), the source of supplemental protein, supplied 14.9, 13.1,25.0,and 22.0% of the total N in the control, urea, starch, and starch plus urea diets, respectively. Intakes and Dlgestlbllltles
Efects
of Urea.
Intake, ruminal digestion,
Chromic oxide was used as a marker to and passage to the duodenum of DM, OM, measure ruminal and total tract digestibilities of starch, ADF, and NDF were not affected by DM, OM, CP, ADF, NDF, and starch. Twice supplementing urea in the diet (Tables 2 and 3). daily, at 0700 and 1900 h, on d 5 through 14 of The daily quantities (kilograms per day) of each period, 10 g of chromic oxide were placed in the rumen via the ruminal cannula. Fecal grab samples were obtained at 0700 and 1900 h during the last 5 d of each period. F e d samJouIIlal of Dairy Science VoL 74, No. 4, 1991
OM, starch, ADF, and NDF digested postnuninally also were not altered by supplementing urea in the diet, but OM digested postruminally expressed either as a percentage of OM intake
EpFE(JTs
1325
OF UREA AND STARCH ON RUMEN FERMENTATION
(P< .07)or as a percentage of OM that passed to the duodenum (P < .@ was increased I) by supplementing urea in the diet (Table 2). The significantly greater percentage of OM (62.8 vs. 57.7%) digested postruminally for diets s u p plemented with urea compared with diets not supplemented with urea can be explained by the concomitant increase in percentage of fiber digested postruminally (Table 3). Digestibilities of ADF (P < .09) and NDF (P < .11) that passed to the small intestine were increased by 5.4 and 5.7 percentage units, respectively, when urea was added to the diet (Table 3). Therefore, small increases in total tract apparent digestibility coefficients were obtained for DM, OM, starch, ADF, and NDF when urea was added to the diet, but these small increases were not significant Vables 2 and 3). Our data support previous reports (8, 46) indicating that DMI of lactating dairy cows was not affected by feeding less than 1.0% of the
total dietary DM as u r a In contrast, Polan et al. (32) depressed DMI of lactating dairy cows when urea was included in the concentrate to supply 1% of the total dietary DM. As we Observed with lactating COWS, ruminal DM digestion was not affected, but postruminal DM disappearance! was significantly improved by infusing 9.8 g/d of urea into the rumen of sheep fed high concentrate diets (31). Likewise, Ortigues et al. (30) reported that ruminal digestion of DM and ADF was not affected by supplementing urea in diets consisting of fescue hay. However, postruminal digestion of DM and ADF was improved when urea was supplemented to the diet, because the quantity of DM and ADF that disappeared from the cecum and colon was increased, whereas the amounts of DM and ADF digested in the small intestine were not altered significantly. Therefore, the i m p v e d postrumjnal digestibilities of DM, OM, ADF, and NDF may have been the result
TABLE 2. Least squares means for intake and digestibility of DM, OM, and starch in various segments of the gastrointestinal tract of lactating cows fed diets that contained fish meal and were supplemented with starch, urea, or starch plus urea Starch
Item
DM h e ,Wd ADl-r.1 96
Control Urea
Starch
+urea
SEM
Urea
Starch
23.1 69.0
23.0 712
21.6 68.0
21.0 71.3
.6 1.7
57 .14
.78
21.0 6.1 29.8 9.8 47.2 14.9 8.7 40.9 57.4 70.6
20.9 5.3 25.6 9.1 44.0 15.6 9.9 472 62.9 72.7
19.6 5.4 8.8 44.9 14.2 8.3 42.1 57.9 70.1
19.1 52 27.6 8.9 46.6 13.9 8.9 46.0 62.7 73.5
5 .7 3.0 .6 2.6 .6 .6 2.3 1.9 1.7
7.6 3.4 45.5 42 4.0 515
7.5 2.6 34.4 4.9 4.7 62.3
8.3 4.1 49.1 4.2 4.0 47.4
8.0 4.2 52.4 3.8 3.7 46.0
2 .4 4.4 A .4 4.1
94.3
94.6 96.7
92.6 96.5
95.4 985
1.3 .7
.02
OM h a ,Wd Apparently digested in the m e n , kg/d Percentage of OM intake Truly digested in the m e n , kg/d Percentage of OM intake Passage to duodenum, kg/d Appanntty digested P o ~ Wd , PCTcentage of OM mtake Percentage of passage to duodenum ADlT,1 %
28.0
58
.14
.02 59 .98 .30 .98 .ll 22 .98 .94 .94
.46 .46
.03 .03
.46 .47
.64 .78 .75 .16
.W .04
Starch
Intake, Wd Apparently digested in the rumen, kg/d Percentage of starch intake Passage to duodenum, kg/d ApP-aY diststed postnmunau ’ Y, kg/d Percentage of starch intake Percentage of starch passage to duodenum ADlT.1 %
97.0
A1
.os
.70
30
55 29
21 .05
26 29
.74 .39
%teraction bestarch and urea was not significant (P > .lo). l~pparentlydigested in thc total g~tmintestinaltract
J o d of Dairy Science Vol. 74. No. 4, 1991
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CAMERON ET AL.
TABLE 3. Least squares means for intake and digestibility of ADP and NDP in various segments of the gastrOinttstiaal tract of lactating cows fed dicts that c o n t a i d fish meal and werc supplemented with starch, urea, or starch plus una. significance of difference (P
