Long-Term Effects of Acetate and Propionate on Voluntary Feed Intake by Midlactation Cows

Long-Term Effects of Acetate and Propionate on Voluntary Feed Intake by Midlactation Cows A. C. SHEPERD and D. K. COMBS Department of Dairy Science, University of Wisconsin, Madison 53706

ABSTRACT Our objective was to separate the effects of physical fill and acetate production in the regulation of voluntary feed intake. Eight ruminally fistulated Holstein cows in midlactation were fed a low forage diet or a high forage diet with or without continuous ruminal infusion of buffered acetate or propionate in a duplicated 4 × 4 Latin square with 21-d periods. Cows consumed about 3.5% of body weight as dry matter, and voluntary dry matter intake (DMI) was approximately 6% greater when cows were fed the low forage diet than when cows were fed the high forage diet. Infusion of 7.1 Mcal of net energy for lactation as acetate or propionate resulted in a reduction in DMI relative to the DMI when the high forage diet was fed alone; propionate infusion reduced intake more than did acetate infusion. Consumption of neutral detergent fiber was approximately 1.19 and 1.25% of body weight when cows were fed the low and high forage diets, respectively. Milk production was approximately 35 kg/d regardless of the diet fed, but an increase in milk fat production by cows receiving the acetate or propionate infusion resulted in an increase in fat-corrected milk. Neither neutral detergent fiber fill nor a threshold for acetate utilization appeared to limit DMI. ( Key words: acetate, propionate, intake) Abbreviation key: HF = high forage diet, iNDF = indigestible NDF, LF = low forage diet, REBW = ruminal empty BW. INTRODUCTION Voluntary feed intake decreases as the proportion of forage in the diet increases. Conrad ( 1 1 ) noted a negative correlation between the amount of forage in the diet and DMI. Because less digestible diets contain larger amounts of indigestible material that must be cleared from the rumen before more feed can be consumed, Conrad ( 1 1 ) attributed the decrease in

Received May 7, 1997. Accepted April 6, 1998. 1998 J Dairy Sci 81:2240–2250

voluntary intake when cows were fed high forage diets ( HF) to a combination of digestibility of the diet, passage of indigestible material from the rumen, and the physical capacity of the cow. Mertens ( 2 5 ) also attributed the decrease in voluntary intake of HF to limitations in physical capacity but found that the amount of NDF in the diet was closely correlated with voluntary intake. Therefore, Mertens ( 2 5 ) proposed that NDF was the factor that limited intake. Although this relationship is true over a wide range of diets, the variability in voluntary intake that results when lactating cows are fed diets varying in NDF content by 5 to 10 units suggests that factors in addition to NDF content regulate intake. As the primary end products of ruminal fermentation, VFA have been studied as possible signals that cause the cessation of eating (4, 5, 15, 18, 30). Although studies suggest that acetate (3, 5, 18, 29) and propionate (5, 15, 30) are involved in short-term regulation, only one study ( 2 9 ) reported voluntary intake when VFA were infused over a period of several weeks. As the proportion of forage in the diet increases, the ratio of acetate to propionate tends to increase because of lesser amounts of propionate that are produced ( 6 ) . However, moles of acetate produced remain relatively constant over a range of diets (1, 14, 17, 33). Studies using lactating cows ( 1 ) have shown that acetate production (moles per day) was similar between cows fed diets containing 10 to 17% hay and cows fed diets containing 38 to 42% hay despite the fact that the ratios of acetate to propionate were 1.4 and 1.8 and the total VFA concentrations were 104 and 83 mmol/L for cows fed the low forage diets ( LF) and HF, respectively. Davis ( 1 4 ) reported that the ratios of acetate to propionate varied from 1.3 to 3.3 but that total acetate production varied by only 1.2 mol/d (28.1 to 29.3 mol/d) when diets contained 13 or 42% hay, respectively. In both experiments (1, 14), feed intake was restricted, which prevented conclusions regarding the effect of acetate on voluntary feed intake. Recent work ( 3 3 ) using steers fed diets containing 20 or 80% alfalfa hay supports the observations of Annison et al. ( 1 ) and Davis ( 1 4 ) (i.e., acetate production remained con-

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stant despite diets that varied widely in the ratio of forage to concentrate). In a recent study with steers (33), feed was offered for ad libitum consumption, and voluntary intake did not differ among diets. The responses of steers, however,may not be indicative of the responses of lactating cows that consume three to four times maintenance requirements. The constancy of acetate production over a wide range of diets suggests that acetate could help to regulate feed intake. However, NDF content of the diet is confounded with acetate production. We hypothesized that the lactating cow can metabolize a specific maximum amount of acetate per day. When acetate production from ruminal degradation of the feed exceeds its capacity to be utilized, DMI is reduced. If HF produce more acetate per kilogram of DM than do LF, the cow theoretically must reduce intake to prevent production of excess acetate. According to the hypothesis, infusion of propionate would provide additional energy without causing a reduction in voluntary feed intake. Physiological mechanisms for such regulatory systems are unclear. The following experiment was conducted to separate the effects of NDF and acetate production on voluntary feed intake by increasing the amount of acetate produced during ruminal fermentation. MATERIALS AND METHODS Eight multiparous Holstein cows were grouped by DIM and assigned to one of two squares in a duplicated 4 × 4 Latin square. To minimize differences in DMI between the squares, cows in square 1 began the experiment 28 d prior to cows in square 2. The DIM for cows in square 1 averaged 95 d and ranged from 84 to 112 d; the DIM for cows in square 2 averaged 88 d and ranged from 69 to 103 d on d 1 of the first period. A standard early lactation diet was fed to all cows from parturition until the beginning of the 14-d preexperimental period during which a diet with a ratio of forage to concentrate of 30:70 was fed. Experimental periods were 21 d in length (14 d of treatment adaptation and 7 d of data collection). Experimental periods were separated by a 7-d period during which all cows received the standard early lactation diet to minimize residual effects of treatments. Recombinant bST was administered to increase milk production and to maintain a high DMI. Cows received half of the recommended dose (12.5 mg) of bST (Posilac; Monsanto Co., St. Louis, MO) weekly starting during wk 9 of lactation ( d 1 of the 14-d preexperimental period) to minimize fluctuations in DMI caused by bST treatment. All procedures involving the cows were approved by the University of

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Wisconsin-Madison College of Agriculture and Life Sciences Institutional Animal Care and Use Committee. The four treatments consisted of a LF, HF, the HF plus infusion of intraruminal acetate (glacial acetic acid; Fisher Scientific, Fair Lawn, NJ), and the HF plus infusion of intraruminal propionate (propionic acid; Aldrich Chemical Co., Inc., Milwaukee, WI). Propionate infusion was included as a treatment to act as a positive control for the additional energy provided by the acetate infusion. In a preliminary experiment conducted to estimate DMI and ruminal pH, the experimental diets were fed to a group of 14 lactating cows, and ruminal fluid was collected using a stomach tube. The acetate or propionate solutions were then prepared to provide the difference in energy intake between the HF and LF. Acids were diluted in 40 L of water ( 3 1 ) and neutralized to match the ruminal pH of cows fed the LF (pH 5.5) using a 3:1 (wt/wt) mixture of NaOH (caustic soda no. 2 flake; Oxychem, Dallas, TX) and KOH (Mallinckrodt Baker, Inc., Phillipsburg, NJ) (35). Feedgrade NaCl and KCl were diluted in 40 L of water and infused into the rumens of cows fed the LF or the HF so that the Na and K intakes were similar between diets. The CP content of the diets was formulated to meet the NRC ( 2 8 ) requirement for undegradable intake protein at the expected level of intake. As a result, HF had a higher CP content than did LF. Composition of the diets is presented in Table 1; composition of the acid solutions is presented in Table 2. The total mixed diets were offered twice daily in amounts to provide 10% orts; water was available at all times. Solutions of acetate or propionate were pumped directly into the rumen using peristaltic pumps (Mec-o-matic VSP-20; W. W. Grainger, Inc., Lincolnshire, IL); acid-resistant tubing was used to deposit the infusate approximately 30 cm from the ruminal fistula (29). Tubing was fused to a weighted and perforated 200-ml plastic bottle to cause mixing of the infusate with ruminal fluid before entering the rumen ( 2 3 ) and to prevent the acid from being deposited on the wall of the rumen. Acid solutions were infused into the rumen for approximately 23 h/d; cows were detached from the pumps only during milking. Cows were adapted to the acid and salt infusion over a 4-d period; 25, 50, 75, and 100% of the acid or salt diluted in 40 L of water were infused on d 1, 2, 3, and 4, respectively, of each period. Intake and milk production were recorded daily throughout the experiment, and samples of the forages and concentrates were taken weekly. Dry matter was determined by drying at 60°C for 48 h in Journal of Dairy Science Vol. 81, No. 8, 1998

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a forced-air oven, and diets were adjusted weekly to account for changes in the DM content of the components. Dried feed samples were ground through a 1-mm screen using a Wiley mill (Arthur H. Thomas, Philadelphia, PA) and analyzed for NDF using aamylase and sodium sulfite (38), for ADF (38), for CP ( 2 ) , and for ash content ( 2 ) . Absolute DM was determined by drying a subsample at 100°C for 24 h. Nutrient composition of the diets was calculated using the composition of the components. Dried and ground (1-mm) dietary components were used to reconstruct the total mixed diets, and rate and extent of ruminal NDF degradation were determined by incubation in vitro for 0, 2, 4, 8, 12, 18, 24, 30, 36, 48, 72, and 96 h with buffer and ruminal fluid from a cow fed HF. Rate was calculated as the natural log of the slope of the amount of NDF remaining at each time

TABLE 1. Formulation and composition of diets1 fed to lactating cows.

Formulation, % of DM Alfalfa silage Corn silage Cracked corn Wheat middlings Corn gluten meal Soybean meal, 44% CP CaHPO4 CaCO3 MgO Trace mineral salt2 Vitamin premix3 Composition, DM basis NDF, % ADF, % CP, % Ca,4 % P,4 % ME,5,6 Mcal/kg NEL,6 Mcal/kg DM, % as fed In vitro NDF degradation Lag, h Extent, % Rate, /h

Preexperimental diet LF

HF

24.0 6.0 28.5 26.9 0 12.6 0.17 0.99 0.23 0.4 0.1

32.0 8.0 24.4 23.1 0 10.8 0.14 0.85 0.20 0.4 0.1

48.0 12.0 8.3 11.6 5.7 13.8 0.02 0 0.2 0.4 0.1

30.2 16.9 18.6 0.81 0.61 2.69 1.63 62.5

31.8 19.7 18.7 0.82 0.58 2.63 1.60 56.8

35.9 25.8 24.8 0.73 0.49 2.51 1.55 48.1

. . . . . . . . .

3.07 45.7 0.056

4.85 42.9 0.05

1Ratios of forage to concentrate: preexperimental diet, 30:70; low forage diet (LF), 40:60; and high forage diet (HF), 60:40. 2Composition: 95% NaCl, 0.34% Fe, 0.005% Co, 0.034% Cu, 0.20% Mn, 0.35% Zn, 0.007% I, and 0.002% Se. 3Provided 2665 IU/g of vitamin A, 900 IU/g of vitamin D, and 0.90 IU/g of vitamin E. 4Determined from a composite sample by the Wisconsin Soil and Plant Analysis Lab (Marshfield). 5Metabolizable energy. 6Calculated from the NRC (28).

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TABLE 2. Infusion parameters for cows fed a low forage diet (LF), a high forage diet (HF), or the HF with continuous ruminal infusions of acetate (HFA) or propionate (HFP). Treatment1

Acetate, mol/d Propionate, mol/d NaOH, mol/d KOH, mol/d NaCl, mol/d KCl, mol/d Infusion volume, L/d Infusion rate,2 ml/min ME3 Infusion,4 Mcal/d

LF

HF

HFA

HFP

0 0 0 0 22.1 8.3 40 29 0

0 0 0 0 22.1 8.3 40 29 0

36.0 0 22.1 8.2 0 0 40 29 7.5

0 20.5 15.6 5.1 6.6 3.0 40 29 7.5

1Ratios of forage to concentrate: LF, 40:60 (31.8% NDF); HF, 60:40 (35.9% NDF). 2Solution was infused for approximately 23 h/d. 3Metabolizable energy. 4Acetate and propionate were assumed to contain 0.2094 and 0.3672 Mcal/mol of ME, respectively.

plotted versus time, and extent was calculated as 100 minus the percentage of NDF remaining at 96 h. Milk samples were taken during consecutive p.m. and a.m. milkings on d 20 and 21 of each period and analyzed for fat, protein, lactose, SCC, and SNF content (Wisconsin DHI Cooperative Center, Appleton). Ruminal fluid was sampled daily 2 h after the p.m. feeding. One hundred milliliters of ruminal fluid were obtained from the anterior dorsal, anterior ventral, medial dorsal, medial ventral, posterior dorsal, and posterior ventral locations within the rumen; composited by cow; and strained through two layers of cheesecloth. A glass electrode was used to measure the pH within 20 min of collection, and duplicate 10-ml subsamples were acidified with 1 ml of 25% (wt/vol) metaphosphoric acid and frozen (–20°C ) for later analysis of VFA by GLC ( 9 ) . Lanthanum oxide ( 2 0 ) was used as a marker to measure total tract digestibility and was dosed at 12-h intervals from d 7 to 21 to provide 0.8 g of La/d. Lithium-Co-EDTA and Cr-mordanted fiber were prepared as described by Ude´n et al. ( 3 6 ) and were used as markers for liquid and solid passage rates, respectively. The Li-Co-EDTA was dried and ground using a mortar and pestle; Cr-mordanted fiber was prepared by mordanting wheat straw NDF and grinding the dried fiber through a 2-mm screen using a Wiley mill. Both Li-Co-EDTA (21 g ) and Cr-mordanted fiber (20 g ) were placed in the rumen at 0800 h on d 18; no attempt was made to manually mix markers with ruminal contents. Fecal grab samples were taken at 0, 6,10, 14, 18, 22, 26, 30, 36, 42, 48, 60, 72, 84, and 96 h after dosing to determine the rate of passage. At

INFUSION OF VOLATILE FATTY ACIDS AND FEED INTAKE

each time an additional 50 g of wet material were composited by cow for digestibility measurements. Samples were dry-ashed (10), and fecal marker concentrations were determined by direct current plasma emissions spectroscopy (Spectra Metrics, Inc., subsidiary of Beckman Instruments, Inc., Andover, MA). Fractional rates of passage were calculated as the slope of the descending portion of the natural logarithm of marker concentration on time. Apparent digestibilities ( A D ) of DM, OM, NDF, and ADF were determined using the equation AD = 100 – (100 × Md/ Mf × Nf/Nd), where Md = concentration of the marker in the diet; Mf = concentration of the marker in the feces, Nf = concentration of the nutrient in the feces, and Nd = concentration of the nutrient in the diet. Infused VFA were assumed to be 100% digestible and were assumed not to affect the digestibility of other components of the diet; DM and OM from infused VFA and salts were included in digestibility calculations. Blood samples were collected from the coccygeal vein into heparinized vacutainer tubes 3 h after the a.m. feeding on d 20 of each period. The samples were immediately placed on ice and transferred within 45 min to NaF-coated centrifuge tubes for the separation of plasma. Aliquots of plasma were stored at –20°C for later analysis of glucose (Sigma procedure 510; Sigma Chemical Co., St. Louis, MO), insulin (Coat-aCount kit TKIN1 with dilution of standards for bovine plasma; Diagnostic Products Corp., Los Angeles, CA), and BHBA (19). Vacutainer tubes containing EDTA were used to collect blood for NEFA ( 2 4 ) analysis and were processed as described previously. Rumens were evacuated, the volume and wet weight of the ruminal contents were determined using precalibrated and tared 70-L barrels, and ruminal empty BW ( REBW) was measured 3 h after the a.m. feeding on d 0 and 21 of each period. The density of ruminal contents was calculated as weight volume. Ten percent of the ruminal contents was separated during the evacuation process, and duplicate 300-g subsamples were dried at 60°C for 72 h, ground through a 1-mm screen using a Wiley mill, and analyzed for NDF, ADF, and CP as described previously. The indigestible NDF ( iNDF) content in the rumen was determined by incubating dried and ground ruminal contents in situ. Neutral detergent fiber remaining after 96 h of incubation was considered to be iNDF. Cows in square 2 were under severe heat stress during the last collection period, resulting in extreme variability in production data; no data from square 2 during period 4 were used in the analysis. Additionally, one cow in square 1 was removed from the

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experiment during period 4 because of an injury unrelated to the experiment. Design and Analysis Data were analyzed as a replicated Latin square. The model was Yijkl = m + Si + Cj( i) + Pk( i) + Tl + STil + Eijkl where m = overall mean, Si = random effect of square ( i = 1 or 2), Cj( i) = random effect of cow within square ( j = 1 to 4), Pk( i) = random effect of period within square ( k = 1 to 4), Tl = fixed effect of treatment ( l = 1 to 4), STil = interaction of square and treatment, and Eijkl = residual. Preplanned orthogonal contrasts were used to determine significance of the main treatment effects of diet and infusions. Contrasts were HF versus LF, HF versus intraruminal VFA infusions, and intraruminal infusions of acetate versus intraruminal infusions of propionate. Significance was declared at P < 0.05 unless otherwise noted. Calculations were completed using PROC GLM of SAS (32). RESULTS AND DISCUSSION Prior to the beginning of the experiment, experimental diets were fed to 14 nonfistulated cows grouped by milk production and DIM; groups averaged 29 kg/d of milk production at 187 DIM and weighed approximately 590 kg. Cows fed the LF and the HF consumed 3.25 and 2.75% of BW as DM, respectively. The difference in NEL intake by cows fed the LF and the HF was approximately 7 Mcal/d (data not shown). Infusate solutions were formulated to provide the expected difference in NEL intake. The metabolizable energy contents of acetate and propionate were assumed to equal 0.2094 and 0.3672 Mcal/ mol, respectively (35); 90% of the metabolizable energy was assumed to be available as NEL. Therefore, the acetate and propionate infusates contained 36.0 and 20.5 mol of acid, respectively, and provided approximately 18% of total NEL requirements. Annison et al. ( 1 ) reported daily acetate production of 37.5 to 47.5 mol/d when lactating cows consumed about 17 kg/d of diets containing 38% hay. Lactating cows fed a 42% hay diet consumed 19 kg of DM and produced about 30 mol/d of acetate (14). Journal of Dairy Science Vol. 81, No. 8, 1998

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TABLE 3. Mineral intake1 by cows fed a low forage diet (LF), a high forage diet (HF), or the HF with continuous ruminal infusions of acetate (HFA) or propionate (HFP). Treatment2 LF Ca, g 216 P, g 153 Na, g 556 K, g 793 Cl, g 1165 S, g 55 DCAD,3 meq/ kg of DM 29

HF

HFA

HFP

188 126 565 962 1228 87

182 122 564 938 144 85

172 115 560 905 477 80

33

138

110

1Includes

dietary sources and minerals in infusate. of forage to concentrate: LF, 40:60 (31.8% NDF); HF, 60:40 (35.9% NDF). 3Dietary cation-anion difference = (percentage of Na/0.0229 + percentage of K/0.0391) – (percentage of Cl/0.003545) + ( 2 × percentage of S/0.03207) ( 8 ) . 2Ratios

Assuming that the acetate production by cows in the current experiment was similar to that of cows in the previous studies (1, 14), the infusion of 36 mol/d markedly increased metabolizable acetate in this experiment. Attempts to infuse unbuffered dilute acid solutions into test animals resulted in the cessation of rumination followed by a rapid decline in ruminal pH and symptoms consistent with metabolic acidosis within 8 h. Partial neutralization of acid solutions was necessary to infuse large amounts of acetate or propionate. All cows received approximately 508 g/d of Na and 320 g/d of K from the infusions (Table 3). Total Na intake (diet plus infusion) was approximately 2.2% of DM, which was greater than the 1.6% recommended by the NRC ( 2 7 ) for lactating dairy cows but less than the 3.7% fed to beef cows by Meyer et al. ( 2 6 ) without adverse effects. Total Cl intake was less than 2.0% of DM, which was below the maximum recommended by the NRC (27). Total K intake was 3.0 and 3.7% of DM for cows fed the LF and the HF, respectively. Beede et al. ( 7 ) reported K intake at 1.2% of DM by heat-stressed cows, but the maximum tolerable concentrations of K in the diets of dairy cows has not been well established (28). Both dietary and infused salts were used to calculate the dietary cation-anion difference ( 8 ) ; values were positive and similar among treatments. Cows consumed about 3.5% of BW as DM (Table 4). Voluntary DMI was approximately 6% greater when cows were fed the LF than when cows were fed the HF. Infusion of 7.1 Mcal of NEL as acetate or propionate resulted in a reduction in DMI relative to Journal of Dairy Science Vol. 81, No. 8, 1998

the DMI when the HF was fed alone. Infusion of propionate had a more inhibitory effect on feed intake than did acetate infusion. Similar to DMI, intakes of NDF and iNDF were reduced when cows were fed the HF relative to the LF. Intakes of NDF were reduced by 3% when acetate was infused and by 9% when propionate was infused. Consumption of NDF was approximately 1.19% of BW when cows were fed the LF and 1.25% of BW when cows were fed the HF. Infusion of acetate and propionate resulted in a decrease in NDF intake as a percentage of BW relative to the NDF intake when HF was fed alone, but means did not differ between infusion treatments. Cows on all treatments exceeded recommended CP and RUP intakes (28). Diets were formulated to contain sufficient CP to meet NRC ( 2 8 ) requirements for undegradable intake protein at the level of intake predicted using Mertens’ model (25). Cows in this experiment consumed more DM than predicted. Additionally, laboratory analysis of the concentrate mix for the HF indicated that the CP content was higher than estimated from the published values used to formulate the diets (28), resulting in a greater difference in CP content between HF and LF than intended. Therefore, cows fed the LF, HF, HF plus acetate infusion, and HF plus propionate infusion consumed approximately 1.3, 2.8, 2.0, and 2.0 kg more CP per d, respectively, than required (28). Intake of iNDF was 4.6 and 5.4 kg/d when cows were fed the LF and HF, respectively. Intakes of iNDF in our experiment were higher than those reported by others ( 1 3 ) when similar diets were fed but were similar as a percentage of BW. Infusions of VFA resulted in a reduction in iNDF of 0.2 kg/d when acetate was infused and 0.5 kg/d when propionate was infused relative to the iNDF content when the HF was fed alone; propionate had a greater effect than did acetate. Intake of iNDF was greater when cows were fed the HF than when cows were fed the LF. Cows consumed 0.66 to 0.77% of BW as iNDF, which did not differ between the LF and HF but was lower with infusion than without infusion and was lower with propionate infusion (0.70%) than with acetate infusion (0.73%). Despite differences among treatments in intake, iNDF content of the rumen was about 4 kg, regardless of treatment. Milk production (Table 4 ) was approximately 35 kg/d and did not differ because of diet or infusion. A 0.28-kg/d increase in milk fat production when acetate was infused and a 0.08-kg/d increase in fat production when propionate was infused resulted in the production of 4.8 and 1.7 kg more 4% FCM by cows receiving acetate or propionate, respectively, compared with the production of FCM by cows receiv-

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ing the HF without VFA infusion. Infusion of acetate resulted in 16% more milk fat and 9% more 4% FCM production than did infusion of propionate. Propionate infusion resulted in milk with a higher percentage of protein than did the infusion of acetate, but daily protein production did not differ among treatments. Milk lactose and SNF contents were unaffected by diet and infusion. Our data support earlier work by Rook and Balch ( 3 1 ) in which infusion of acetate resulted in an increase in milk production; however, unlike the earlier experiment, cows in our experiment responded to propionate infusion by increasing milk production. The REBW was approximately 625 kg and did not differ among treatments (Table 4). Change in REBW during the 21-d period was similar (about 0.25 kg/d) when cows were fed the LF and the HF. Infusion of acetate or propionate resulted in more than a 0.5 kg/d increase in REBW gain relative to the HF fed alone; acetate and propionate produced similar amounts of REBW gain.

Ruminal pH was about 5.6 when cows were fed the LF and 6.0 when cows were fed the HF (Table 5). Infusion of either acetate or propionate resulted in an increase in ruminal pH relative to the pH when HF was fed alone. That the ruminal pH was higher than the pH of the infused solutions indicated that the cow produced more buffer than was necessary to buffer that acid produced by fermentation of the diet. Ruminal pH did not differ between infusion treatments, and the 0.2-unit increase in pH with infusions was not expected to have a significant effect on the results of the experiment. Concentrations of VFA in ruminal fluid (Table 5 ) were about 154 mM when cows were fed the LF and the HF and 183 mM when acetate or propionate was infused. Although higher than expected, total VFA content was similar to data recently reported by Dado and Allen (12, 13) using diets similar in NDF content and digestibility. Profiles of VFA reflected differences in the treatments in agreement with data reported by Ørskov et al. (29). Infusion of acetate caused an

TABLE 4. Voluntary feed intake, milk production, milk composition, and BW of cows fed a low forage diet (LF), a high forage diet (HF), or the HF with continuous ruminal infusions of acetate (HFA) or propionate (HFP). Contrast Treatment1 LF

HF

HFA

HFP

SEM

LF vs.HF

HF vs. HF Plus infusion

HFA vs. HFP

P Voluntary intake, kg/d DM OM NDF iNDF2 ADF CP Production, kg/d Milk 4% FCM Fat Protein Lactose SNF Milk composition, % Fat Protein Lactose SNF BW, kg Empty BW, kg Empty BW gain, kg/d Voluntary intake, % of BW DM NDF iNDF 1Ratios

26.3 24.1 8.4 4.6 5.0 4.9

25.7 23.3 9.2 5.4 6.4 6.3

24.9 22.5 8.9 5.2 6.3 6.1

23.5 21.3 8.4 4.9 5.9 5.8

0.4 0.4 0.2 0.1 0.1 0.1

0.002 0.001 0.022 0.037 <0.001 <0.001

0.005 0.007 0.014 0.012 0.005 0.020

0.014 0.016 0.048 <0.001 0.004 0.098

34.9 33.5 1.23 1.15 1.65 3.04

34.9 32.8 1.18 1.15 1.67 3.07

36.2 37.6 1.46 1.13 1.77 3.15

36.2 34.5 1.26 1.20 1.78 3.23

1.2 1.1 0.05 0.03 0.06 0.10

0.461 0.199 0.137 0.780 0.175 0.301

0.347 0.026 0.007 0.765 0.188 0.301

0.995 0.042 0.004 0.148 0.840 0.512

3.55 3.34 4.69 8.73 700 621 0.31

3.41 3.34 4.78 8.81 702 623 0.20

4.08 3.16 4.87 8.73 716 629 0.73

3.58 3.37 4.89 8.96 708 628 0.76

0.09 0.07 0.07 0.11 4.5 3.8 0.20

0.145 0.467 0.055 0.343 0.086 0.130 0.237

0.002 0.347 0.225 0.790 0.082 0.259 0.034

0.001 0.022 0.822 0.096 0.162 0.853 0.894

3.76 1.19 0.66

3.66 1.31 0.77

3.47 1.24 0.73

3.32 1.19 0.70

0.05 0.02 0.01

<0.001 0.039 0.068

<0.001 0.002 0.002

0.016 0.080 <0.001

of forage to concentrate: LF, 40:60 (31.8% NDF); HF, 60:40 (35.9% NDF). NDF.

2Indigestible

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TABLE 5. Effects of a low forage diet (LF), a high forage diet (HF), or the HF with continuous ruminal infusions of acetate (HFA) or propionate (HFP) on ruminal pH and VFA concentrations. Contrast Treatment1 LF Ruminal pH VFA, mM Acetate Propionate Isobutyrate Butyrate Isovalerate Valerate Total VFA, mM VFA, mol/100 mol Acetate ( A ) Propionate ( P ) Isobutyrate Butyrate Isovalerate Valerate A:P 1Ratios

HF

HFA

HFP

SEM

LF vs. HF

HF vs. HF Plus HFA vs. infusion HFP

5.6

5.9

6.1

6.0

<0.1

<0.001

P 0.040

0.339

84.2 39.9 1.7 19.6 3.7 4.9 154.4

87.6 33.2 3.0 19.0 4.4 5.6 153.1

119.9 32.9 3.7 20.2 4.6 5.5 185.1

89.6 61.0 2.4 19.3 5.2 6.3 182.4

3.9 4.1 0.2 0.5 0.2 0.3 5.2

0.003 0.551 <0.001 0.890 <0.001 0.023 0.004

0.002 0.013 0.548 0.219 0.018 0.405 <0.001

<0.001 <0.001 <0.001 0.192 0.013 0.071 0.676

2.4 1.4 0.1 0.5 0.2 0.2 0.14

0.246 0.274 <0.001 0.047 0.028 0.452 0.004

0.778 0.032 0.060 0.025 0.280 0.156 0.989

<0.001 <0.001 <0.001 0.640 0.094 0.109 <0.001

54.5 25.8 1.1 12.7 2.4 3.2 2.14

56.8 21.7 2.0 12.5 2.9 3.7 2.67

65.2 17.8 2.1 11.1 2.6 3.0 3.70

50.0 33.1 1.4 10.8 2.9 3.5 1.65

of forage to concentrate: LF, 40:60 (31.8% NDF); HF, 60:40 (35.9% NDF).

increase in acetate concentration without changing propionate concentration. Similarly, propionate infusion increased propionate concentration without altering acetate concentration. Ruminal fluid of cows fed the LF contained about 55% acetate and 26% propionate, and the ruminal fluid of cows fed the HF contained about 57% acetate and 22% propionate. Ratios of acetate to propionate were 2.1 and 2.7 when cows were fed the LF and the HF, respectively; infusion of acetate resulted in a ratio of acetate to propionate of 3.7, and propionate infusion resulted in a ratio of 1.7. Ruminal digesta characteristics are shown in Table 6. Dry matter percentage was higher when cows were fed the LF than when cows were fed the HF and was higher when cows were fed the HF without VFA infusions than when cows were fed the HF with VFA infusions. Ruminal OM, NDF, and ADF contents did not differ among treatments. Ruminal contents weighed about 80 kg (11% of BW) and did not differ among treatments. Ruminal volume was 83 L in cows fed the LF, the HF, and the HF plus propionate infusion; ruminal volume was about 10% greater in cows receiving the HF plus acetate infusion. Density of ruminal contents and mass of DM and NDF in the rumen were unaffected by treatment. Ruminal volume was lower than that reported by others, even though our cows were larger; however, the mass of both wet digesta and DM was greater Journal of Dairy Science Vol. 81, No. 8, 1998

(12, 13, 21). We did not find differences between the HF and the LF unlike Dado and Allen ( 1 2 ) who reported such a difference, and we did not find the reduction in volume and mass with VFA infusion that others (12, 20) have reported with the addition of inert bulk. Our data indicated that the reduction in feed intake caused by VFA infusion was unrelated to changes in ruminal size or composition of the ruminal contents. The fractional rate of passage of Cr-mordanted fiber from the rumen ranged from 0.068 to 0.078/h, and mean retention time ranged from 12.9 to 16.1 h (Table 7). Passage of the liquid phase marker ranged from 0.093 to 0.105/h; mean retention times ranged from 9.8 to 10.9 h. Passage of the liquid and solid phase markers was slightly higher than reported by others (22), probably a factor of the increased ruminal osmolality that resulted from the infusion of salts or VFA ( 3 7 ) and the small particle size of the Crmordanted fiber. Neither passage rate nor mean retention time differed among treatments. In contrast to data reported by others (29), digestibility of DM was higher when cows received infusions of acetate or propionate than when cows were fed the HF without infused VFA, but digestibility of DM did not differ between VFA infusions. Digestibility of NDF was similar between the LF and the HF but was 17% higher with the infusion of acetate or propionate. Digestibility of ADF was also improved by the infu-

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INFUSION OF VOLATILE FATTY ACIDS AND FEED INTAKE

TABLE 6. Ruminal digesta characteristics of cows fed a low forage diet (LF), a high forage diet (HF), or the HF with continuous ruminal infusions of acetate (HFA) or propionate (HFP). Contrast Treatment1 LF

HF

HFA

HFP

SEM

LF vs. HF

HF vs. HF Plus HFA vs. infusion HFP P

Digesta composition DM, % OM, % of DM NDF, % of DM ADF, % of DM Ruminal mass, kg Wet digesta DM OM NDF ADF iNDF2 Ruminal volume, L Density, kg/L 1Ratios

15.5 90.3 63.5 42.3

14.6 90.1 62.2 43.7

13.4 89.6 63.0 44.6

13.1 89.3 60.0 42.6

0.5 0.5 3.9 3.2

0.005 0.199 0.660 0.169

0.043 0.284 0.865 0.914

0.564 0.609 0.521 0.101

79.6 12.3 11.1 7.8 5.2 4.1 82.5 0.97

78.8 11.5 10.4 7.1 5.0 3.9 82.3 0.96

86.9 11.7 10.5 7.4 5.2 4.2 93.0 0.95

79.7 10.4 9.3 6.2 4.5 3.8 83.3 0.96

3.0 0.5 0.5 0.5 0.5

0.486 0.059 0.045 0.082 0.285 0.453 0.283 0.566

0.209 0.458 0.391 0.570 0.590 0.806 0.151 0.974

0.076 0.074 0.065 0.076 0.044 0.160 0.035 0.857

3.4 0.03

of forage to concentrate: LF, 40:60 (31.8% NDF); HF, 60:40 (35.9% NDF). NDF.

2Indigestible

sions relative to the digestibility of ADF when HF was fed alone; infusions of acetate or propionate affected NDF and ADF digestibility similarly. Reasons for the increase in DM and fiber digestibility without changes in passage rate associated with VFA infusions are unclear. Plasma glucose, insulin, NEFA, and BHBA concentrations (Table 8 ) reflected the expected differences in VFA metabolism. Glucose concentration was

higher with the infusion of propionate than with the infusion of acetate but was unaffected by forage concentration in the diet or VFA infusion relative to the HF fed alone. These data supported work by others ( 3 4 ) that showed that ruminal propionate was used as a glucose precursor. The NEFA content of plasma was used as an indicator of energy balance among treatments (16). Low NEFA concentrations supported calculations of NEL balance and indicated that

TABLE 7. Effects of a low forage diet (LF), a high forage diet (HF), or the HF with continuous ruminal infusions of acetate (HFA) or propionate (HFP) on ruminal rate of passage and mean retention time of solids and liquids and total tract digestibility of nutrients. Contrast Treatment1 LF

HF

HFA

HFP

SEM

LF vs. HF

HF vs. HF Plus HFA vs. infusion HFP P

Rate of passage, /h Cr-Mordanted fiber Li-Co-EDTA Mean retention time, h Cr-Mordanted fiber Li-Co-EDTA Nutrient digestibility,2 % DM OM NDF ADF 1Ratios

0.070 0.093

0.068 0.105

0.075 0.095

0.078 0.099

0.007 0.006

0.678 0.941

0.296 0.372

0.713 0.237

14.7 10.9

16.1 9.8

15.6 10.7

12.9 10.2

1.8 0.5

0.264 0.208

0.233 0.284

0.543 0.407

67.1 68.5 45.5 43.8

63.8 65.1 42.2 42.6

67.0 68.2 48.2 48.8

69.1 70.3 50.4 51.3

1.4 1.4 1.9 2.7

0.716 0.592 0.452 0.185

0.021 0.022 0.007 0.032

0.241 0.238 0.359 0.440

of forage to concentrate: LF, 40:60 (31.8% NDF); HF, 60:40 (35.9% NDF). calculations included DM and OM from diets and infusions.

2Digestibility

Journal of Dairy Science Vol. 81, No. 8, 1998

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SHEPERD AND COMBS TABLE 8. Plasma glucose, insulin, NEFA, and BHBA concentrations of cows fed a low forage diet (LF), a high forage diet (HF), or the HF with continuous ruminal infusions of acetate (HFA) or propionate (HFP). Contrast Treatment1 LF

HF

HFA

HFP

SEM

LF vs. HF

71.20 11.48 106.42 8.77

68.93 9.06 128.97 8.35

66.66 11.48 137.42 13.81

74.66 14.33 108.28 6.97

2.44 1.91 20.55 1.17

0.648 0.876 0.378 0.432

HF vs. HF Plus infusion 0.538 0.099 0.794 0.148

HFA vs. HFP

P Glucose, mg/dl Insulin, ng/dl NEFA, meq/L BHBA, mg/dl 1Ratios

0.020 0.235 0.258 <0.001

of forage to concentrate: LF, 40:60 (31.8% NDF); HF, 60:40 (35.9% NDF).

cows were in a positive energy balance; there were no differences in NEFA concentrations among treatments. Ruminal infusion of acetate resulted in a twofold increase in BHBA concentration in plasma relative to the infusion of propionate. Plasma BHBA was about 8.5 mg/dl when cows were fed the LF or the HF and was slightly lower ( 7 mg/dl) when cows received the HF plus propionate infusion. The increase in BHBA suggested that acetate increased ketone body formation as in other studies (29), but clinical signs of ketosis were not observed in the current experiment. Plasma measurements supported production parameters; both VFA were absorbed from the rumen, but acetate was apparently used primarily for synthesis of milk fat, and propionate was used as a glucose precursor. Some propionate was converted to protein and excreted in milk. Both VFA treatments resulted in BW gain. Cows consumed more DM than expected, resulting in a voluntary NEL intake that was approximately 2.3 Mcal/d less when the HF was fed than when the LF was fed (Table 9). Therefore, the addition of the 7.1 Mcal of NEL/d as either acetate or propionate increased total NEL intake to 108 and 103% of LF, respectively. Dietary NEL intake was higher when cows were fed the LF than when cows were fed the HF, was higher when cows were fed the HF than when cows were fed the HF plus either infusion, and was higher when cows were infused with acetate than when cows were infused with propionate. Total NEL intake did not differ when the LF was compared with the HF. Infusion of acetate or propionate increased total NEL intake by approximately 10% relative to the HF fed alone; infusion of acetate resulted in a total intake of 2.1 Mcal of NEL/d more than infusion of propionate. Utilization of NEL for milk production reflects the increase in 4% FCM production with both infusions Journal of Dairy Science Vol. 81, No. 8, 1998

relative to the HF fed alone and with acetate versus propionate. Energy for maintenance did not differ among treatments, and energy for REBW gain was higher with the VFA infusions than without them. Therefore, total NEL utilization was higher when cows received infusions of acetate or propionate than when cows were fed the HF alone but did not differ by diet or infusion. Greater than 90% of energy intake could be accounted for by milk production, main-

TABLE 9. Calculated values for energy partitioning of cows fed a low forage diet (LF), a high forage diet (HF), or the HF with continuous ruminal infusions of acetate (HFA) or propionate (HFP). Treatment1

NEL Intake, Mcal/d Total2 Diet3 Infusion4 NEL Utilization, Mcal/d Total5 Milk6 Maintenance7 REBW Gain8 Recovery,9 %

LF

HF

HFA

HFP

42.1 42.1 0

39.8 39.8 0

45.5 38.4 7.1

43.4 36.3 7.1

38.1 24.8 11.7 1.6 90.4

37.0 24.3 11.7 1.0 93.3

43.5 27.8 11.9 3.7 96.1

41.2 25.5 11.8 3.9 94.7

1Ratios of forage to concentrate: LF, 40:60 (31.8% NDF); HF, 60:40 (35.9% NDF). 2Total NE intake from diet and infusion. L 3Voluntary NE intake. L 4Content of NE from infusions (conversion of metabolizable L energy to NEL of acetate and propionate was assumed to be 95%). 5Sum of NE for milk, maintenance, and ruminal empty BW L (REBW) gain. 6Calculated as 0.74 × kilograms of 4% FCM. 7Calculated as 0.086 × BW0.75. 8Calculated as 5.12 × kilograms of REBW gain/d. 9Percentage of total NE intake accounted for as milk, mainL tenance, and REBW gain.

INFUSION OF VOLATILE FATTY ACIDS AND FEED INTAKE

tenance, and REBW gain using prediction equations published by the NRC (28). The infusion of energy in the form of acetate or propionate supplied up to 8% more NEL than cows voluntarily consumed. This energy was used to support an increase in 4% FCM production and increased BW gain. The supplementation of additional energy as acetate or propionate resulted in a decrease in voluntary intake but not enough to compensate for the energy provided by the VFA. Results of this experiment show that cows consumed more DM than was predicted by either ruminal fill or NEL requirements (25). The LF was formulated to meet NEL requirements so that ruminal fill would not be limiting; the HF was formulated to be limiting by ruminal fill as predicted by NDF capacity (1.2% of BW) of the cow. Cows fed the LF were expected to consume 26.4 kg of DM based on an NDF intake of 1.2% of BW or 23.8 kg of DM to meet their 38.1-Mcal requirement for NEL. Actual DMI exceeded expected DMI by 1 and 11% when based on NDF and NEL, respectively. Similarly, cows that received the HF consumed approximately the same amount of DM as that predicted by NDF but approximately 6% more DM than that predicted from requirements for NEL when the requirement for NEL was reduced to account for 7.1 Mcal from the infusions. CONCLUSIONS Data did not support the hypothesis that ruminal production of acetate, not ruminal fill, regulates the feed intake of dairy cows. Apparently, ruminal fill and energy absorption are not the only factors that regulate intake. The form of absorbable energy has a significant impact on intake. In this experiment, cows fed HF utilized more infused energy as acetate than as propionate. The consumption of more NDF than expected and the utilization of more energy than voluntarily consumed together suggest that lactating cows do not rely entirely on physical fill or energy content of the diet to regulate feed intake. Also, cows do not appear to be nearing a threshold for acetate or propionate metabolism when fed normal lactation diets. Additionally, the observation that cows partitioned the extra infused energy to milk fat and BW gain discourages the assumption that a genetic potential for milk production drives feed intake; the form of the energy supplied also clearly affects utilization. The amount of additional energy that the cow is able to utilize is unknown and may depend on the availability of other nutrients. We speculate that cows utilized the additional energy provided as acetate or

2249

propionate until another nutrient, perhaps an amino acid, became limiting. Regulation of voluntary feed intake by lactating dairy cows is a complex process involving not only NDF content of the diet and energy requirements for milk production but also the form of the energy supplied and factors that affect the partitioning of energy. REFERENCES 1 Annison, E. F., R. Bickerstaffe, and J. L. Linzell. 1974. Glucose and fatty acid metabolism in cows producing milk of low fat content. J. Agric. Sci. (Camb.) 82:87–95. 2 Association of Official Analytical Chemists. 1980. Official Methods of Analysis. 12th ed. AOAC, Washington, DC. 3 Baile, C. A., and C. L. McLaughlin. 1987. Mechanisms controlling feed intake in ruminants: a review. J. Anim. Sci. 64: 915–922. 4 Baile, C. A., and W. H. Pfander. 1966. A possible chemosensitive regulatory mechanism of ovine feed intake. Am. J. Physiol. 210:1243–1248. 5 Battacharya, A. N., and M. Alulu. 1975. Appetite and insulinmetabolite harmony in portal blood of sheep fed high or low roughage diet with or without intraruminal infusion of VFA. J. Anim. Sci. 41:225–233. 6 Bauman, D. E., C. L. Davis, and H. F. Bucholtz. 1971. Propionate production in the rumen of cows fed either control or highgrain, low-forage diet. J. Dairy Sci. 54:1282–1287. 7 Beede, D. K., W. K. Sanchez, and C. Wang. 1992. Macrominerals. Pages 272–286 in Large Dairy Herd Management. H. H. Van Horn and C. J. Wilcox, ed. Am. Dairy Sci. Assoc., Champaign, IL. 8 Block, E. 1984. Manipulating dietary anions and cations for prepartum dairy cows to reduce incidence of milk fever. J. Dairy Sci. 67:2939–2948. 9 Brotz, P. G., and D. M. Schaefer. 1987. Simultaneous determination of lactic and volatile fatty acids in microbial fermentation extracts by gas-liquid chromatography. J. Microbiol. Methods 6:139–144. 10 Combs, D. K., and L. D. Satter. 1992. Determination of markers in digesta and feces by direct current plasma emission spectroscopy. J. Dairy Sci. 75:2176–2184. 11 Conrad, H. R. 1966. Symposium on factors influencing the voluntary intake of herbage by ruminants: physiological and physical factors limiting feed intake. J. Anim. Sci. 25:227–235. 12 Dado, R. G., and M. S. Allen. 1995. Intake limitations, feeding behavior, and rumen function of cows challenged with rumen fill from dietary fiber or inert bulk. J. Dairy Sci. 78:118–133. 13 Dado, R. G., and M. S. Allen. 1996. Enhanced intake and production of cows offered ensiled alfalfa with higher neutral detergent fiber digestibility. J. Dairy Sci. 79:418–428. 14 Davis, C. L. 1967. Acetate production in the rumen of cows fed either control or low-fiber, high-grain diets. J. Dairy Sci. 50: 1621–1625. 15 Deetz, L. E., and P. J. Wangsness. 1981. Influence of intrajugular administration of insulin, glucagon, and propionate on voluntary feed intake of sheep. J. Anim. Sci. 53:427–433. 16 Dunshea, F. R., A. W. Bell, and T. E. Trigg. 1989. Relations between plasma non-esterified fatty acid metabolism and body fat mobilization in primiparous lactating goats. Br. J. Nutr. 62: 51–61. 17 Esdale, W. J., G. A. Broderick, and L. D. Satter. 1968. Measurement of ruminal volatile fatty acid production from alfalfa hay or corn silage rations using a continuous infusion isotope dilution technique. J. Dairy Sci. 51:1823–1830. 18 Forbes, J. M., J. N. Mbanya, and M. H. Anil. 1992. Effects of intraruminal infusions of sodium acetate and sodium chloride on silage intake by lactating cows. Appetite 19:293–301. Journal of Dairy Science Vol. 81, No. 8, 1998

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